101.
Identification Of Genetic Variants In The Low Density Lipoprotein Receptor Gene Causing Familial Hypercholesterolemia And Its Sequence Homology With Mus Musculus
by Ameer Hassan (2014-VA-504) | Dr. Wasim Shehzad | Dr. Muhammad Yasir Zahoor | Dr. Muhammad Tayyab.
Material type: ; Literary form:
not fiction
Publisher: 2016Dissertation note: Familial hypercholesterolemia (FH) is caused by mutations in the genes coding for the low-density lipoprotein receptor (LDLR), apolipoprotein B-100, or proprotein convertase subtilisin/kexin type 9 (PCSK9). In this study, a molecular analysis of LDLR gene was performed in 20 unrelated patients from Pakistan. All patients were clinically diagnosed with definite or possible hypercholesterolemia according to a uniform protocol and internationally accepted WHO criteria. Preferable study was made to highlight the Genetic variation in Exon 4 of LDLR gene associated with defective catabolism of cholesterol effecting lipid metabolism which results in Familial Hypercholesterolemia. The extraction of genomic DNA was done from all selected blood samples. By selecting primers they were synthesized and optimized on extracted DNA samples. PCR product was sequenced and aligned. Mutations in the LDLR gene and its sequenced homology with Mus musculus were analyzed. We didn’t found any polymorphisms in the LDLR gene exon 4. So we concluded that there is no association between SNPs and increased levels of cholesterol in Pakistani population. More research should be carried out in Pakistan by increasing the sample size and considering the other regions of LDLR gene. This study will help the early detection and treatment of such cases and may ultimately reduce the incidence of mortality due to myocardial infarction. Apart from diagnosis, we also suggest it will be a potential therapeutic strategy to manage FH. Availability: Items available for loan: UVAS Library [Call number: 2538-T] (1).
102.
Molecular Investigation Of Low Density Lipoprotein Receptor Gene Causing Familial Hypercholesterolemia And Its Evolutionary Relationship With Pan Troglodytes
by Rida Zainab (2014-VA-808) | Dr. Maryam Javed | Dr. Asif Nadeem | Prof. Dr. Tahir Yaqub.
Material type: ; Literary form:
not fiction
Publisher: 2016Dissertation note: Familial Hypercholesterolemia (FH) phenotype is related to improper metabolism of low density lipoproteins due to mutations in Low-density lipoprotein receptor (LDLR) gene with increased risk of ischemic heart disease. Genetic variants in LDLR gene are associated with defective catabolism of cholesterol effecting lipid metabolism which results in familial hypercholesterolemia. It occurs in both forms: Homozygous Familial Hypercholesterolemia and Heterozygous Familial Hypercholesterolemia.
Patients having high cholesterol were identified by observing the values of their serum lipid profile test reports. Their detailed history was taken and blood samples from the identified patients of familial hypercholesterolemia were collected. DNA extraction was done by Organic method. Primers were synthesized and PCR was conducted using optimized recipe and conditions. PCR products were sequenced.
Sequenced data was analyzed using Chromas or BioEdit software. BLAST was performed and sequences were aligned individually by comparing it to the reference sequence. This showed difference in any specific position of a mutated sequence against the reference sequence. CLUSTALW aligned all the sequences together in one time. Sequences were compared with reference sequence to detect the presence of any mutation or SNPs.
SNPs were identified manually and the peaks were observed in order to determine if the genotype is heterozygous or homozygous. Statistical Analysis was done and any amino acid change due to the observed SNPs was determined by using Expasy Translate Tool. It was found that both the SNPs showed amino acid changes. In the end, homology analysis was done which showed that Homo sapiens had their LDLR gene closest to that of Gorilla gorilla gorilla. Availability: Items available for loan: UVAS Library [Call number: 2551-T] (1).
103.
Polymorphism Analysis Within Tata-Box Of Bovine Lactoferrin Gene And Its Association With Mastitis In Sahiwal Cows
by Kashmala Haroon (2014-VA-04) | Dr. Sehrish Firyal | Dr. Immad Rashid | Dr. Muhammad Tayyab.
Material type: ; Literary form:
not fiction
Publisher: 2016Dissertation note: Mastitis is one of the most important diseases in dairy cows throughout the world, and is responsible for significant economic losses to the dairy industry especially in Pakistan. Several factors are responsible for this disease and about 20% bovines are suffering with this disease. Mastitis susceptibility and resistance is influenced by genetic variation of animals. Variations to polymorphisms in LF gene assume critical part of the immune response to mastitis. Polymorphism within LF gene may influence immune response to the mastitis in bovines. Recent study shows that promoter region of LF gene is highly polymorphic among bovines.
Present study was planned to identify polymorphism analysis within TATA-box of bovine LF gene and its association with mastitis. Multiple blood samples were collected from Sahiwal cows having clinical and sub-clinical mastitis. 10 samples were collected as a control. DNA extraction was done by organic extraction method and then quantification was done by Nanodrop. Amplification and sequencing was performed to get desire sequence of the gene. Comparative study of obtained sequence results were analyzed by using NCBI blast. Bioinformatics analysis was done with the help CLUSTAL W and BioEdit softwares.
Two novels and one reported SNPs were discovered within TATA-box of LF gene that might be having strong genetic association with mastitis in Sahiwal cows. This gene is strong candidate gene to differentiate between mastitis susceptible and resistant Sahiwal cows.
Availability: Items available for loan: UVAS Library [Call number: 2584-T] (1).
104.
Polymorphism Analysis Of Exon 2,5 And 10 Of Bovine Lactoferrin Gene And Its Association Within Mastitis In Sahiwal Cows
by Sidra Mukhtar (2014-VA-223) | Dr. Sehrish Firyal | Dr. Sultan Ali | Dr. Muhammad Wasim | Dr. Muhammad Avais.
Material type: ; Literary form:
not fiction
Publisher: 2016Dissertation note: A few factors militate against understanding the milk production capability of bovines. Mastitis is the most costly and the prevalent production-limiting disease of dairy animals in Pakistan and somewhere else on the world. Susceptibility and resistance to mastitis is a complex characteristic and impacted by hereditary variation of animals. Among these variations, the polymorphisms in LF assumes critical part in the immune responses to mastitis.
Susceptibility and resistant to mastitis is a complex trait and influenced by genetic variation of the immunity genes of the animals. Among these variations, polymorphism in Lactoferrin gene (LF) play important role in immune responses to mastitis. Polymorphism in exons 2, 5 and 10 of Lactoferrin gene are associated with mastitis susceptibility and resistance. It is a potential candidate gene for screening of the mastitis susceptible and resistant dairy cows.
The present study was designed for the identification of polymorphism in LF gene associated with mastitis. Blood samples from 20 Sahiwal cows having clinical and subclinical mastitis were sampled. Blood samples of 10 normal Sahiwal cows were also collected. DNA was extracted. Specific primers for amplification of LF gene were designed by using Primer 3 software.
LF gene was amplified and sequenced to get the full length sequence of the gene. Comparative analysis of resulted sequences was done with the help of NCBI BLAST. Multiple sequence alignment was done by using CLUSTAL W and BioEdit softwares. Protein analysis was done with ExPasy translate tool and the development of 3D structure were using PYMOL software.
Availability: Items available for loan: UVAS Library [Call number: 2582-T] (1).
105.
Sequence Analysis Of Mitochondrial Atpase 8/6 Gene Variants In Sindh Ibex (Capra Aegagrus Blythi)
by Javeria Zafar (2014-VA-222) | Dr. Asif Nadeem | Dr. Maryam Javed | Prof. Dr. Abu Saeed Hashmi.
Material type: ; Literary form:
not fiction
Publisher: 2016Dissertation note: ATPase 8/6 gene plays a vital role in survival of an organism by generating energy in the form of ATP synthase. Considering the importance of ATPase8/6 gene in energy generating, present study has been designed to characterize this gene in Sindh ibex. The characterization of ATPase8/6 gene might be helpful for deriving phylogenetic relationship among different species and identifying new functions among the related species. Tissue/blood samples (n=15) were collected from Kirthar National Park, Sindh. Standard DNA extraction method was used for DNA extraction. PCR primers were designed by Primer3 software and amplification of gene was done by Polymerase Chain Reaction. PCR product was sequenced bi-directionally by Big Dye TM Terminator on ABI 3130XL Genetic Analyzer. Multiple sequence alignment was performed for polymorphism identification. Genetic diversity was calculated by using DNAsp. Phylogenetic analysis using the MEGA6 software package and an equally weighted maximum parsimony analysis was performed using the close-neighbor-interchange algorithm. The results indicated that Sindh ibex ATPase8/6 gene was highly similar to Capra caucasica. The results of this data might be helpful in designing effective conservation strategies of different species of wild animal. Availability: Items available for loan: UVAS Library [Call number: 2587-T] (1).
106.
Principles of Cell Biology
by Plopper, George.
Edition: 2nd ed.Material type: ; Literary form:
not fiction
Publisher: USA: Jones & Bartlett; 2016Availability: Items available for loan: UVAS Library [Call number: 571.6 Plopper 31735 2nd 2016 Genetics] (1).
107.
Development Of Novel Mtdna Metabarcodes For The Species Differentiation Of Class Aves
by Syeda Rida Mehak Sherazi (2010-VA-477) | Dr. Muhammad Imran | Dr. M. Yasir Zahoor | Mr. Shahid Abbas.
Material type: ; Literary form:
not fiction
Publisher: 2016Dissertation note: The Folmer COI mtDNA universal primers that are considered standard for DNA barcoding of life contain so many mismatches against the target sequences of vertebrate origin that they often end in failure to amplify many of vertebrate DNA extractions. This discrepancy favors for the selection and designing of new metabarcode primers that can be used to identify all individuals of vertebrates or at least all individuals represented in a class of Vertebrata such as Class Aves. The current study embarks on such an endeavor. In this study development of new mtDNA metabarcode (16SrRNA) that can be used as universal primers to amplify almost all species of Class Aves for different forensic and molecular biodiversity analyses.
Blood/feather/tissue samples were collected from Class Aves (one specimen from every order reported to be present in Pakistan). DNA was extracted from the collected specimens through standard organic method, qualified and quantified and then PCR-amplified using novel universal primers selected from aligned mtDNA sequences originating from all Aves mitochondrial DNA genomes submitted to different online sequence databases such as NCBI nucleotide database. The sensitivity of PCR also be assessed using a range of DNA concentrations. The amplified products were sequenced on ABI Genetic Analyzer following Sanger’s dideoxy method of sequencing. The correctness of obtained mtDNA sequences were examined visually in Chromas Lite 2.1 software and then alignment of these sequences were performed against highly similar DNA sequences in NCBI nucleotide databases using BLAST in order to identify origin of unknown mtDNA sequences. With the help of sequencing and phylogenetic studies specificity of the universal primer set confirmed and presented as a novel metabarcode (16SrRNA) for species level identification of large number of Avian species
In summary, we present universal method for species classification of Aves using a targeted parallel sequencing approach. Both sequencing and phylogenetic studies experiments confirm
Summary
82
specificity of universal primer set. Although promising results were obtained with current settings, rapid improvement of bench top instruments will further develop method with less hands-on, fewer sequencing errors and lower detection limit. So, in future, this barcode can be used for species identification in various fields of study such as meat adulteration, illegal trade, food mislabeling and molecular estimation of biodiversity. Availability: Items available for loan: UVAS Library [Call number: 2617-T] (1).
108.
Identification And Expression Analysis Of Genes Involved In Obsessive Compulsive Disorder In Pakistani Population
by Javeria (2008-VA-627) | Prof. Dr. Masroor Ellahi Babar | Dr. Muhammad Wasim | Prof. Dr. Muhammad Abdullah.
Material type: ; Literary form:
not fiction
Publisher: 2016Dissertation note: The background of this study is that WHO reports that psychiatry disorders affect worldwide 0.8 to 2% population. Anxiety illnesses are a class of illness associated with unreasonable and disturbing sensation of fear and tension. There are several types of anxiety disorders, such as panic disorder, agoraphobia, specific phobia, social phobia, OCD. Obsessive-compulsive disorder is a chronic disabling condition. OCD is characterized by repetitive, intrusive thoughts, images, and impulses and by repetitive, ritualistic physical or mental acts performed to reduce the attendant anxiety. The severity of OCD depends on the amount obsessions and compulsions. The Yale-Brown Obsessive Compulsive Scale (Y-BOCS) is a reliable and consistent scoring system that can be used to categorize OCD. The major genes involve in OCD are SLC6A4, BDNF, SLC1A1 and COMT genes.
The study was enrolled patients treated for OCD. Blood samples have been collected from the patients. DNA extracted from fresh blood. Primers were designed. Then DNA amplification have done by Bio-Rad thermal cycler. Then gel electrophoresis was done for PCR product quantification. PCR products precipitated and sequenced. SNPs were identified. Real-time quantitative RT-PCR was performed for each sample with TaqMan Universal PCR mastermix which showed down regulation of COMT gene in OCD patients in Pakistani population.
The aim of this study was SNP identification in Pakistani Population in Obsessive Compulsive disorder and to analyze the gene expression of COMT gene involved in OCD in Pakistani Population.
Availability: Items available for loan: UVAS Library [Call number: 2620-T] (1).
109.
Development Of Novel MtDNA Metabarcodes For Species Differentiation Of Class Reptilia
by Imran Tariq (2014-VA-505) | Dr. Muhammad Imran | Dr. Wasim Shehzad | Dr. Saadat Ali.
Material type: ; Literary form:
not fiction
Publisher: 2016Dissertation note: The FoImer COI: mtDNA. universal primers that are considered standard for DNA barcoding of life contain so many =matches against the target sequences of vertebrate men tat they often end in failure to amplify many of vertebrate DNA eurections. This disaepancy fawn foe the seaman and designing of new metabarcode panes that can be used m ideally all inditdrals of vertebrees or at least all individuals represented in a class of Vertebrate such as Cass Reprilia. The current study was embadang on such an endeavor
The proposed study was develop new m5DNA membarc ode that may be used as universal Kilns; to amplify almost all species of Class Repalia for different formic and molectdr biodivesity analyses.
Blood and tissue samples were collected from Class Repdha (at :east 24 species from every ceder reported to be present in Pakistan) DNA was extracted from the collected specimen through stacdasd organic method. qualified and =meted and then PCR-amplified using novel universal primers selected from aligned =DNA sequences origtadng from all repdlian mitochondria DNA pnomes submitted to diens online sequence databases such as NCB: micleotide database. Tne sensitiviry. of PCR was assessed using a range of DNA come:madam. The amplified products were sequenced on A131 Genetic Analyzed following Sarge's dideacy method of sequencing.
The correctness of obtained croDNA sequences were examined visually in Chromes Lite 2.1 software and then alipmmt of these sequences were per: waxed agitinc highly similar DNA sequences in NCBI nu6eonde databases using BLAST in order to identify the coigin of la-noun =DNA sequences
sequencing everimeas and phyla...net< studies was confirm the specificity of the universal primer set developed and present a novel metabarcode for species level identification of large number of reptelian species. So, In future this barcode can be used for species identification in various fields of study such as illegal trade and molecular estimation of boidiversity.
Availability: Items available for loan: UVAS Library [Call number: 2628-T] (1).
110.
Homology & Polymorphism Analysis Of Cc2d1a Gene In Human And Canine For Cognitive Function
by Hafiz Qamar Abbas (2014-VA-214) | Dr. Muhammad Yasir Zahoor | Dr. Wasim Shehzad | Dr. Saadat Ali.
Material type: ; Literary form:
not fiction
Publisher: 2016Dissertation note: Cognitive disability is a group of genetically heterogeneous abnormality that leads to variable degrees of cognition deficits. It has been shown that inherited disorders can be caused by mutations in large number of different genes and there is evidence for the presence of as yet unknown genes in a significant proportion of patients. This disease can affect 1-3% of overall population and higher in consanguineous families. We aimed to identifying the homology and polymorphism of the gene CC2D1A between human and canines. The present research work was carried out in four phases. The first phase was including enrolment of 10 affected non relevant families with disease history and consent was taken on consent forms as approved by IRB, UVAS. Secondly DNA extraction was done by using standard lab protocols. Thirdly amplification of the selected domains of selected gene (CC2D1A) was done through PCR amplification after designing primers of the selected domains. Sequencing of the amplified products has to be done through Sanger method and mutation analysis was conducted for variants We found two new asynonymous mutation one is deletion of c. 1664_1664delA which lead to the change in the normal function of protein (88%) and other is heterozygous mutation c.1921A/T that result in amino acid change from R to W (12%). Whereas homology analysis shows that deletion region is partially conserved as it code different amino acid but some key domains are conserved. This homology shows that deletion in this region can change the protein expression which can relate to unconscious condition like behavioral or mental retardation. This will be helpful in providing genetic counseling services to indigenous population for intellectual disability cases. Availability: Items available for loan: UVAS Library [Call number: 2627-T] (1).
111.
Introduction to Biotechnology / 3rd ed.
by Thieman, William J | Palladino, Michael A.
Material type: ; Literary form:
not fiction
Publisher: India: Pearson Education, 2014Availability: Items available for loan: UVAS Library [Call number: 660.6 Thieman 31793 2nd 2014 Biotechnology] (5).
112.
Biochemistry, Molecular Biology And Genetics / 6th ed.
by Lieberman, Michael A.
Edition: 6th ed.Material type: ; Literary form:
not fiction
Publisher: New Delhi: Wolters & Kluwers; 2014Availability: Items available for loan: UVAS Library [Call number: 572.8076 Lieberman 31789 6th 2014 Biochemistry] (1).
113.
Calculations in Molecular Biology and Biotechnology : A Guide to Mathematics in the Laboratory / 2nd ed.
by Stephenson, Frank H.
Edition: 2nd ed.Material type: ; Literary form:
not fiction
Publisher: India: Academic Press; 2014Availability: Items available for loan: UVAS Library [Call number: 572.80151 Stephenson 31847 2nd 2014 Biotechnology] (1).
114.
Molecular Characterization Of Canine Babesiosis In Ticks And Dogs
by Tahira Sarwar (2014-VA-523) | Dr. Wasim Shehzad | Dr. M. Yasir Zahoor | Dr. Ali Ahmed Sheikh.
Material type: ; Literary form:
not fiction
Publisher: 2016Dissertation note: Babesia canis is an intra-erythrocytic parasite which cause canine babesiosis in both animals and humans. Currently, there are three sub-species of Babesia canis has been identified i.e Babesia canis canis , Babesia canis vogeli and Babesia canis rossi. Currently used diagnostic methods are clinical symptoms, peripheral blood smear microscopy and serological tests (IFAT and ELISA).Current study was conducted to compare the specificity and sensitivity of blood smear microscopy and PCR techniques to diagnose canine babesiosis. This study is comparative as well as developmental in nature. Although peripheral blood smear microscopy is cost effective and quick method of diagnosis in case of high or moderate parasitaemia in blood. But the limitations associated with microscopy include false negative diagnosis in case of low parasitaemia in chronic and asymptomatic infection,morphological similarity of Babesia with other species of Plasmodium and Theileria these limitations may lead to misdiagnose the infection due to which disease may remain unnoticed.Total 50 samples comprising of 25 blood samples and 25 ticks were collected randomly from infected dogs from June, 2015 to November, 2015. These samples were screened microscopically as well as with PCR. Out of 50 samples of dogs and ticks, 18 samples found to be positive for the Babesia canis. 11 samples are Babesia canis vogeli and 07 samples are Babesia canis canis were to be identified in positive samples of dogs and ticks.The results obtained from the study clearly show that PCR is more reliable, precise and sensitive assay for diagnosis of canine babesiosis then microscopy. It is hoped that proposed method to diagnose babesiosis will help to nullify the problems associated with microscopy. This will ultimately facilitate in the formulation of effective treatment control and vaccine development strategies which may eradicate babesiosis. Availability: Items available for loan: UVAS Library [Call number: 2642-T] (1).
115.
Genetic Identification And Characterization Of Pakistani Birds Of Perdicinae Subfamily (Partridge) Through Dna Barcoding Method
by Asim Iqbal Jutt (2013-VA-557) | Dr. Ali Raza Awan | Dr. Muhammad Wasim | Prof. Dr. Kamran Ashraf.
Material type: ; Literary form:
not fiction
Publisher: 2016Dissertation note: Pakistani birds of Perdicinae sub family are cage and game birds. Birds includes Altectoris chukar, Ammoperdix heyi, Ammoperdix griseogularis, Francolinus francolinus and Francolins pondicerianus. Traditional methods of identification were based on the phenotypical characterization of birds, which may lead to incorrect identification, so there was need to explore their characters at DNA level for accurate identification and to establish a DNA reference.
Birds of sub-family Perdicinae have not been genetically characterized in Pakistan. A new precise method “DNA barcoding” was applied using COI gene of mDNA for authentic identification and classification of these birds. Blood and tissue samples of five species (fifteen samples) were obtained. DNA of each sample was extracted by organic method. Amplification of CO1 gene was done by using a universal set of primers BIRDF1, BIRDR1. Sequence of 450bp were analyzed using bioinformatics softwares. Each sample was aligned with its reference sequence of COI gene available on NCBI. Every nucleotide position which did not align with the reference sequence was studied to identify SNPs. A common phylogenetic tree of all partridges showed that they have common ancestor about 0.7 million year ago, F.francolinus, F.pondicerianus and A.heyi shared a common clade whereas A.chukar made a separate clade from the ancestor. A.heyi and F.pondicerianus showed closed resemblance. It has been proved that DNA barcoding is an efficient and accurate molecular tool for species identification and phylogenetic implication. This study established a DNA Data Bank that helped scientists to investigate the biodiversity, taxonomic classification, species identification and also established foundations for molecular biologists to study taxonomic uncertainties at sub species level using SNP based identifying marker. Availability: Items available for loan: UVAS Library [Call number: 2714-T] (1).
116.
Molecular Biotechnology
by Dehlinger, Carolyn.
Edition: 1st ed.Material type: ; Literary form:
not fiction
Publisher: USA: Jones and Bartlett; 2016Availability: Items available for loan: UVAS Library [Call number: 572.838 Dehlinger 32226 1st 2016 Biotechnology] (1).
117.
Statistical Methods in Molecular Evolution
by Nielsen, Rasmus.
Edition: 1st ed. Material type: ; Literary form:
not fiction
Publisher: New Delhi: Springer; 2005Availability: Items available for loan: UVAS Library [Call number: 576.8 Nielsen 32281 1st 2005 Genetics] (1).
118.
Cell and Molecular Biology : Concepts and Experiments / 6th ed
by Karp, Gerald.
Edition: 6th ed.Material type: ; Format:
print
; Literary form:
not fiction
Publisher: New York : John Wiley & Sons, 2010Availability: Items available for loan: UVAS Library [Call number: 575.1 Karp 32259 6th 2010 Genetics] (1).
119.
Molecular Data Analysis Using R
by Ortutay, Csaba | Ortutay, Zsuzsanna.
Edition: 1st ed.Material type: ; Literary form:
not fiction
Publisher: Singapore: Wiley Blackwell; 2017Availability: Items available for loan: UVAS Library [Call number: 572.33 Ortutay 32257 1st 2017 Statistics] (1).
120.
Molecular Biology / 4th ed.
by Weaver, Robert Franklin.
Edition: 4th ed. Material type: ; Literary form:
not fiction
Publisher: USA: McGraw Hill; 2008Availability: Items available for loan: UVAS Library [Call number: 572.8 Weaver 32245 4th 2008 Genetics] (1).
121.
Introduction to molecular immunology/ 1st ed.
by Alfred Ninsonoff.
Edition: 1st ed.Material type: ; Literary form:
not fiction
Publisher: USA: 1982Availability: Items available for loan: UVAS Library [Call number: 616.079 Alfred 12373 1st 1982 Molecular.Biology] (1).
122.
Snp Genotyping Of Cacna1a Gene Implicated In Childhood Absence Epilepsy (Cae)
by Wajeeha Tariq (2010-VA-487) | Dr. Muhammad Wasim | Dr. Ali Raza Awan | Dr. Muhammad tayyab.
Material type: ; Literary form:
not fiction
Publisher: 2016Dissertation note: Childhood absence epilepsy (CAE) is more pediatric epileptic syndrome. It is about 5 to 15% of all childhood epilepsies. CAE is polygenic and multifactorial syndrome. Many different genes other than CACNA1A gene are involved to cause the CAE collectively. Mutation in P/Q type alpha 1 A subunit channel (Cav2.1) gene CACNA1A, leading to the reduction of Cav2.1 activity in both neurons and in expression system. Reduction in Cav2.1 channel activity altered the neurotransmitter release at neocortical synapses. Molecular genetics techniques have identified various mutation in the genes of ion channels such (CACNA1A, CACNA1G, CACNA1H, CACNB4), sodium channel genes (SCN1A, SCN1B and SCN2A) and GABA receptor genes (GABRD and GABRG2). CACNA1A ion channels are the standard mediator of neurotransmission in Central nervous system (CNS) and mutations in this gene play significant role in the generation of absence seizures. Pore forming alpha 1 a (Cav2.1) channels encoded by CACNA1A gene and are usually located in presynaptic neuron.
Present study was aimed to examine coding regions of CACNA1A gene for analyzing the mutations involve in epilepsy.
Blood samples (n = 40) of true CAE representatives were collected from Children hospital Lahore. DNA was isolated from all blood samples through standard organic method. Amplification of CACNA1A gene exon 36 regions was done with specially designed primers.
Later on, results were analyzed through sequencing of target region. Sequenced samples were analyzed through BioEdit software and alignment was done through Clustal Omega software.
It has been identified that absence epileptic patients of Pakistan showed Mutation in exon 36 of CACNA1A gene at position 281258bp and 281285bp which alter the protein sequence. Due to frame shift mutation a stop codon was detected at position 1813 in protein sequence. So a truncated and loss of function Cav2.1 channel might be formed. In epileptic patients, mutation is responsible for the absence seizures.
In the conclusion, we can say that additional study with large number sample is required to amend the effects of these mutations and their associated factors are precisely and perfectly identified. Further, there is need to investigate the other gene variation causing epilepsy in the local population of Punjab Pakistan. This study will ultimately help to develop genetic counseling strategies, gene therapies and parental diagnostic procedures for the Pakistani population.
Availability: Items available for loan: UVAS Library [Call number: 2746-T] (1).
123.
Comparative Genomic Study of Motor Neuron Disease in Horses and Human
by Shakeela Daud (2011-VA-534) | Dr. Muhammad Wasim | Prof. Dr. Tahir Yaqub | Dr. Abu Saeed Hashmi.
Material type: ; Literary form:
not fiction
Publisher: 2017Dissertation note: cd not submitted Availability: Items available for loan: UVAS Library [Call number: 2810-T] (1).
124.
Exploration Of Genetic Polymorphisms And Differential Expression Analysis Of Bovine Alpha-Lactalbumin And Osteopontin Genes Involved In Milk Composition
by Sidra Manzoor (2010-VA-92) | Dr. Asif Nadeem | Muhammad Imran | Dr. Abu Saeed Hashmi.
Material type: Publisher: 2017Dissertation note: Economically important traits of dairy animals are usually controlled by a large number of genes. The identification of the single nucleotide polymorphisms in potential genes has been associated with economically important traits. During lactation, mammary epithelial cells produced large amounts of specific milk proteins. Due to the expression sites, physiological properties and chromosomal localization, LALBA and SPP1 genes might be considered as candidate genes for milk composition in buffalo. Alpha-lactalbumin (LALBA) gene has been reported to be highly transcribed in transition and peak phase while late lactation exhibited its decline with progressive rise in SPP1 expression. This project was designed to investigate the effect of single nucleotide polymorphism that influencing the gene expression thus modulates the milk protein content in Nili Ravi. Samples of unrelated Nili-Ravi buffalo were collected from two Government, Buffalo Research Institute, Pattoki, and Livestock Production and Research Institute (LPRI) Bahadarnagar Okara, livestock farms. Milk samples were collected at 15, 90 and 250 days lactation for expression analysis. The genomic DNA was extracted by using the standard Phenol Chloroform Isoamyl alcohol (PCI) protocol. Specific set of primers was designed for the amplification of the LALBA and SPP1 genes. The amplified PCR products were sequenced for the identification of SNPs. To determine the differential expression of bovine LALBA and SPP1 genes, RNA was isolated from milk samples using the TRIzol reagent and converted it into cDNA. Taqman probes were used that are specifically designed to detect and target the DNA sequence. Five intronic polym orphic sites were identified in LALBA while exonic regions exhibited a complete homology with reference sequence. Additionally, eleven polymorphisms were identified in bovine SPP1 gene, six were in coding region and five were
Summary
122
found in intronic portion of the gene. The analysis and correlation of all identified polymorphism was done by using SNPs data analysis software “SNPator”. Results obtained from expression study was stored in in-build software of Real Time PCR and Cycle threshold (Ct) values of LALBA and SPP1 mRNA were compared in individuals of Nili-Ravi buffalo to determine the variation in expression levels. The LALBA gene expression was observed highest in transition phase with a gradual decrease of expression in mid and late lactation. The sample, NR-5, was observed highly expressed (79.30) while NR-2 with low expression (19.28) for alpha lactalbumin in early lactation. The change in LALBA regulation at same stage was considered due to genetic variation of the respective animal. While the SPP1 gene expression was observed with the highest values in peak lactation and remains elevated in late lactation. NR-4 has the highest (72.27) expression among all mastitis free healthy animals while NR-2 was observed with low expression. Thus, the identified SNPs might be used as genetic marker for milk production traits. Gene expression patterns may also help us to understand the molecular mechanisms of bovine LALBA and SPP1 genes influencing milk composition. However, the expression of both genes was considered in a correlation with other genes involved in milk production pathway. Also, the mutational effects of other milk proteins might be involved in determining the expression pattern of both genes in selected animals. Therefore, further studies are likely to explore the regulation of milk protein genes and their translational efficiency during the course of lactation in dairy animals. Availability: Items available for loan: UVAS Library [Call number: 2830-T] (1).
125.
Development Of Novel Mtdna Metabarcodes For The Species Differentiation Of Class Amphibia
by Rehmatullah (2011-VA-365) | Dr. Muhammad Imran | Dr. M. Yasir Zahoor | Dr. Amjad Riaz.
Material type: Publisher: 2017Dissertation note: The Folmer COI mtDNA universal primers that are considered standard for DNA
barcoding of life contain so many mismatches against the target sequences of vertebrate origin
that they often end in failure to amplify many of vertebrate DNA extractions. This discrepancy
favors for the selection and designing of new metabarcode primers that can be used to identify all
individuals of vertebrates or at least all individuals represented in a class of Vertebrata such as
Class Amphibia. The current study embarks on such an endeavor. In this study development of
new mtDNA metabarcode (16SrRNA) that can be used as universal primers to amplify almost all
species of Class Amphibia for different forensic and molecular biodiversity analyses.
Tissue samples were collected from order Urodela of Class Amphibia (Toads , Bull frog
and skittering frogs sample were collected from Punjab, Pakistan). DNA was extracted from the
collected specimens through standard organic method, qualified and quantified and then PCRamplified
using novel universal primers selected from aligned mtDNA sequences originating
from order Urodela mitochondrial DNA genomes submitted to different online sequence
databases such as NCBI nucleotide database. The sensitivity of PCR also be assessed using a
range of DNA concentrations. The amplified products were sequenced on ABI Genetic Analyzer
following Sanger’s dideoxy method of sequencing. The correctness of obtained mtDNA
sequences were examined visually in Chromas Lite 2.1 software and then alignment of these
sequences were performed against highly similar DNA sequences in NCBI nucleotide databases
using BLAST in order to identify origin of unknown mtDNA sequences. With the help of
sequencing and phylogenetic studies specificity of the universal primer set confirmed and
Summary
67
presented as a novel metabarcode (16SrRNA) for species level identification of large number of
Amphibian species.
In summary, we present universal method for species classification of Amphibia using a targeted
parallel sequencing approach. Both sequencing and phylogenetic studies experiments confirm
specificity of universal primer set. Although promising results were obtained with current
settings, rapid improvement of bench top instruments will further develop method with less
hands-on, fewer sequencing errors and lower detection limit. So, in future, this barcode can be
used for species identification in various fields of study such as meat adulteration, illegal trade,
food mislabeling and molecular estimation of biodiversity. Availability: Items available for loan: UVAS Library [Call number: 2874-T] (1).
126.
Molecular Diagnosis Of Brucella Zoonosis As Blood Transfusion Hazard In Metropolitan Population Of Lahore And Okara
by Amna Azam (2011-Va-3560 | Dr. Wasim Shehzad | Dr. Iahtasham Khan | Dr. Muhammad Imran | Dr. Imran Rashid.
Material type: Publisher: 2017Dissertation note: Brucellosis and Coxiellosis are one of the most spreading zoonotic diseases. They both are facultative, intracellular, Gram negative and involved in bioterrorism and agro-terrorism attacks. Brucella abortus and Brucella melitensisare common among all specie of Brucella. Total two hundred Human Blood transfusion samples were collected from hospitals of Okara and Lahore. Blood was collected in vacutainers (without EDTA). After serum isolation serological test RBPT were performed of all samples. Eighty nine out of two hundred were positive to RBPT with seroprevalence of 44.5% (95% Confidence Interval [CI]: 37.61 – 51.4). DNA extraction was done. The concentration of DNA was analyzed through Nanodrop 2000. Then these samples were subjected to Genus specific Real-time PCR analysis. Forty one out of two hundred were positive to Genus specific Real-time PCR with seroprevalence of 20.5% (95% Confidence Interval [CI]: 14.9 – 26.09). Brucella genus positive samples were subjected to two species specific PCR Brucella abortus and Brucella melitensis respectively. Forty one out of forty one Brucella genuspositive samples were positive to Brucella melitensisand none of the sample was positive to Brucella abortus. These two hundred DNA samples were then subjected to Coxiella specific Real-time PCR analysis and 4 were found positive with seroprevalence of 2% (95% Confidence Interval [CI]: 0.06 – 3.94). Results were recorded in the form of Ctvalue. Results indicate that Real-time PCR is more efficient than RBPT and due to increasing seroprevalence in Blood transfusion samples its screening should be included in normal blood transfusion screening procedure through collaboration with Government to prevent transfusion transmitted infections (TTI’s). Availability: Items available for loan: UVAS Library [Call number: 2873-T] (1).
127.
Study Of Arginine Vasopressin (Avp) As A Candidate Gene For Evaluating Silent Estrus Behavior In Nili-Ravi Buffalo
by Muhammad Danish Ahmad (2011-VA-464) | Dr.Maryam Javed | Dr. Asif Nadeem | Dr.Muhammad Zubair Shabir.
Material type: Publisher: 2017Dissertation note: Buffalo is a major contributing animal in livestock and in Pakistan as an agrarian country its economy. Nili- Ravi buffalo also known as “Black Gold of Pakistan” has a high potential of productivity. But its production is often effected by certain reproductive disorders, out of which silent estrus behavior act as a major limiting factor for its production, as it is difficult to proper diagnose of silent estrus it result in low fertility and ultimately yield as low productivity in buffalo. There are a number of reasons involve in silent estrus behavior such as nutrition, environment and genetics. Estrus is a polygenic trait and according to a report about 269 genes are involve in estrus. One of the major effecting gene on estrus is Arginine Vasopressin (AVP), produced by hypothalamus and released by posterior pituitary lobe. Along with a number of role in body it influences the Social behavior neural network, located in limbic system and produce the responses like sexual arousal, partner pairing, mating process and social dominancy. So the AVP accounted as a potential candidate gene for study the silent estrus behavior in Nili-Ravi buffalo. The basic aim to conduct the present study was to identify the Single Nucleotide Polymorphism (SNP) in exonic region of AVP gene and find their association to the silent estrus trait. Fifty samples in blood form of Nili-Ravi Buffalo was taken from B-block research forms UVAS, Ravi campus and Buffalo Research Institute (BRI) Pattoki. DNA was extracted using Phenol Chloroform Iso-amyl alcohol (PCI) based method, then DNA was subjected to PCR amplification, Product was precipitated and sequenced for genetic analysis. To identify the SNPs in obtained sequence the Bioinformatics tools such as BLAST and CHROMAS were applied. The three exonic regions of AVP gene were amplified using site specific primer sets. A total of 6
Summary
58
polymorphic sites were identified, those all were present in exon 1. The bioinformatics analysis using PopGene32 software was performed to analyze the association of identified SNPs to the Silent Estrus Behavior. SNP were analyzed for their effect on trait and one SNP in exon-1 was analyzed for its effect on subjected trait. This genetic characterization of AVP gene may serve as the genetic source for the development of DNA based markers for used in selection of animals with better estrus trait in studies, research and commercial purposes. Availability: Items available for loan: UVAS Library [Call number: 2871-T] (1).
128.
A Thesis Submitted In The Partial Fulfillment Of The Requirements For The Degree
by Ayesha Saddiqa (2011-VA-367) | Dr. Asif Nadeem | Dr. Maryam Javed | Dr. Muhammad Zubair Shabbir.
Material type: ; Format:
print
; Literary form:
not fiction
Publisher: 2017Dissertation note: AIDS epidemic is increasing rapidly in the Eastern Mediterranean Region. Quoting fresh authorized figures collected by the Punjab health department were 97,000 to 125,000 HIV positive people in Pakistan. Number of patients with HIV/AIDS rapidly increased in Punjab. 310 HIV/AIDS cases (35 women and 13 children) have been stated in Punjab in 2014. CCR5 gene is associated HIV infection. Mutation in this gene delayed the progression towards AIDS. In this study blood samples were collected from the laboratory of Punjab Aids Control Program (PACP), primary and secondary health care department, Government of Punjab. Genomic DNA was extracted by using the using FavorPrepTM Blood/Cultured Cell Genomic DNA Extraction Mini Kit. Specific set of primers were designed for the amplification of the targeted gene. The amplified PCR products were precipitated and sequenced for the identification of polymorphisms. Bidirectional sequencing was done for result confirmation. Alignments of sequences were done with the help of NCBI BLAST. Chromas software, Clustal W, UCSC, Bio Edit and SNPedia and Mega 6.0 was used to compile this study. CCR5 32 base pairs allele deletion was found absent in all HIV positive and negative individuals. So, susceptibility of human immuno-deficiency virus type one infection is high in Pakistani population. Genomic comparison was done with non-human primates. Alignment result showed human CCR5 gene homology, 95%, 99%, 94% and 94% with Maccaca mulata (Rhesus Monkey), Pan troglodytes (chimpanzee), Cercocebus atys (sooty mangabey) and Rhinopithecus bieti (black snub-nosed monkey) respectively. So, this homology analysis showed that these non-human primates can be used for development of therapeutic strategies related to human immune deficiency virus. Availability: Items available for loan: UVAS Library [Call number: 2872-T] (1).
129.
Mutational Analysis Of Atp7b Gene Responsible For Wilson’s Disease And Its Homology Analysis In Primates And Mouse
by Amama Ghaffar (2011-VA-375) | Dr. M. Yasir Zahoor | Prof. Dr. Huma Arshad Cheema | Dr. M. Imran | Dr. Amjad Riaz.
Material type: ; Literary form:
not fiction
Publisher: 2017Dissertation note: Copper being an essential element to carry out different cellular processes normally is
maintained through proper regulation mechanisms to avoid its accumulation in the body. ATP7B
gene that codes for ATP dependent P type ATP7B protein controls the regulation of copper in
the body. It is required for the proper delivery of copper to apoceruloplasmin and its excretion
through bile in the form of feces. Therefore, mutation occurring in the ATP7B gene can cause
excessive cellular copper accumulation which results into Wilson’s disease. Variation in ATP7B
gene related to copper transportation leads to Wilson’s disease and transmitted in generation
through recessive pattern of inheritance.
For this study blood samples of fifteen Wilson’s disease affected patients along with
normal individuals of the same family were collected from Children's Hospital & Institute of
Child Health, Lahore. DNA was extracted from blood through organic extraction method
followed by DNA quantification. Amplification of exons 8, 13, 14 and 18 of ATP7B gene was
performed after designing specific primers for these specific regions. Sequencing of amplified
products was done through dideoxy chain termination method. A disease causing mutation of
ATP7B gene c.3155 C>T; p1052 Proline (CCC) to Leucine (CTC) has been mapped on exon 14
in family with Wilson’s disease. This mutation can be used for genetic testing, prenatal
diagnosis and genetic counseling. No mutation was found in exons 8, 13 and 18 which mean that
further study needs to be done to find more local mutation(s) that can be used for fast direct
genetic testing of Wilson’s disease patients or the carriers with heterozygotic conditions who can
develop this disease at any age of their life.
Results
87
MUSCLE and Clustal Omega were used for homology analysis of ATP7B gene
nucleotide and protein sequences that revealed Gorilla to be closest to human regarding coding
sequences, while Clustal Omega output file showed all the species varied highly in their protein
structure homologies. Through the prediction of secondary structure homologies it was seen that
marmoset was closest to humans.
This study helped in providing prenatal diagnosis and genetic screening services in the
country. It has facilitated in selecting animal models for further study and research on ATP7B
gene and molecular pathogenesis of the Wilson’s disease leading to prevention and cure of
disease. Availability: Items available for loan: UVAS Library [Call number: 2892-T] (1).
130.
Mutation Detection Of Cacnb4 Gene Involved In Childhood Absence Epilepsy And Its Comparative Genomics In Mice
by Ayesha Amin (2015-VA-1048) | Dr. Muhammad Wasim | Dr. Sehrish Firyal | Prof. Dr. Aftab Ahmad Anjum.
Material type: ; Literary form:
not fiction
Publisher: 2017Dissertation note: Childhood absence epilepsy (CAE) is due to multiple factors affecting almost 9-12 years children. Depolarization of ion channel activates the channel. CACNB4 gene is affected by epileptic seizures. Disturbance in ion channel can affects different genes as CACNA1G, CACNA1H, CACNA1A, SCN1B, SCN1A,SCN2A and GABA receptor genes. CACNB4 gene has a major role in influencing epilepsy in human.
In present study,it is directed to analyze the mutations in epilepsy present in coding region of CACNB4 gene.
Collection of blood samples were from Children Hospital, Lahore, Punjab Pakistan from CAE patients of epilepsy. By using standard DNA extraction method, DNA was extracted from samples. Primers were designed for the amplification of exon 3 and 13 of CACNB4 gene.
Results were examined after sequencing the samples. BioEdit software was used to study the samples thoroughly. NCBI BLAST was used to align the sequences.
It is investigated that the sequences of CAE patients of epilepsy of CACNB4 gene has mutation at position position 258023bp which changes A>G. In protein sequence, the mutation is at position 413 which changes L (Leucine) to L (Leucine). This mutation has no effect because this is a synonymous mutationwhere the codon CUG is changes to CUA, both codes for same amino acid that is leucine, so no effect at all by this change in exon 13.
Three mutations are present in the intronic region of exon 13 first, second and third at positions 258184bp A deleted, 258289bp and 258191bp of CACNB4 gene respectively. These all mutations are present in intronic region so has no effect in phenotypes of individual.
In conclusion, maximum numbers of samples were needed to observe the effect of mutations and factors that causes epilepsy. This study will now help the researchers to investigate genetic therapies, strategies of genetic counseling and parental diagnosis for the population of Pakistan.
Availability: Items available for loan: UVAS Library [Call number: 2924-T] (1).
131.
Study On Polymorphism Of Promoter Region Of Bovine Lactoferrin Gene And Its Relation With Mastitis In Nili Ravi Buffalo
by Muhammad Asim (2012-VA-636) | Dr. Sehrish Firyal | Dr. Muhammad Yasir Zahoor | Dr. Muhammad Tayyab.
Material type: ; Literary form:
not fiction
Publisher: 2017Dissertation note: Dairy animals in Pakistan and worldwide are facing most persistent and economically affecting
disease mastitis. Only a healthy buffalo can produce good quality milk of physiologically normal
composition. Mammary gland inflammation due to mastitis badly affects the quantity and quality
of milk and this causes big loss to dairy industry. Even province Punjab bears economic losses of
240 million per annum due to mastitis.
Susceptibility and resistance to mastitis is affected by the variation in immunity genes. Among
immunity genes Lactoferrin (LF) have important role in immune defense system and perform
antibacterial, antiviral and anti-inflammatory function. LF is found in most of body fluids like,
milk, blood, tear, saliva, bile and mucous. Polymorphism in promoter region of Lactoferrin gene
is associated with mastitis susceptibility and resistance. For screening of the mastitis
susceptibility and resistance of dairy buffaloes, LF is a potential candidate gene.
The present study was designed for the identification of polymorphism in LF gene associated
with mastitis. Blood samples from 20 Nili Ravi buffalos having clinical and subclinical mastitis
were selected. Blood samples of normal Nili Ravi buffalos were also collected. DNA was
extracted; specific primers for amplification of LF gene were designed with Primer-3 software
by using already reported sequence on NCBI.
Amplification of LF gene performed by PCR and sequenced the amplicon. A comparative
analysis of sequence result was performed by using NCBI BLAST. BioEdit software was used to
perform multiple sequence alignment. Comparison analysis of LF gene promoter region shows
multiple mutations in in clinical and subclinical as compare to reported sequence (accession no.
EF650854) at NCBI. While normal samples sequence results are similar to reference data. These
results show LF is a candidate gene for mastitis resistance. Availability: Items available for loan: UVAS Library [Call number: 2923-T] (1).
132.
Mutational Analysis Of Cacna1ggene Implicated In Childhood Absence Epilepsy And Its Comparative Genomics In Mice
by Fiza Idrees (2015-VA-803) | Dr. Muhammad Wasim | Dr. Ali Raza Awan | Prof. Dr. Aftab Ahmed Anjum.
Material type: ; Literary form:
not fiction
Publisher: 2017Dissertation note: Childhood absence epilepsy (CAE) is the subtype of Idiopathic generalized epilepsy (IGE). It accounts for 2-8% of patients with epilepsy. The frequency of CAE is more in girls than boys. The percentage of CAE in youngsters is 10-12%. In addition to CACNA1G, many other genes can be the possible cause of CAE. The pattern of inheritance of CAE is polygenic and complex. SNP might be a gain of function mutation in T- channel genes that results in increase T-type calcium channel activity. Ion channel genes and genes for GABA receptors are affected in epilepsy. By using various techniques of molecular genetics mutations have detected in genes of calcium channels (CACNA1H,CACNA1I, CACNA1A, CACNA1G and CACNB4), in genes of sodium channels like (SCN1B, SCN2A, SCN1A ) and genes for GABA receptor (GABRG2 and GABRD ). Gain of function mutation in CACNA1G gene and increased activity of α1G channels are the possible reason for abnormal SWD in absence epilepsy.
Aim of this study was to assess acknowledged and/or the novel mutations in CACNA1G gene obtained from local childhood epileptic patients.
Blood samples (n=20) were obtained from CAE patients. These samples were collected from children hospital Lahore. Organic method was used to extract DNA from these collected samples. Specific primers were designed for exon 13 and 17 and these exonic regions were amplified using PCR.
After PCR, sequencing of PCR products was performed and then sequencing results were analyzed using chromas lite software.
It has been observed that CACNA1G gene has two mutations in exon 17. It was noticed that protein sequence was altered and the positions of mutations were 38594bp and at 38635bp 38594bp and at 38635 bp. So SNP was detected and there was a gain of function mutation α1G channel activity. In conclusion, these mutations are responsible for absence seizures in CAE patients.
So, it can be concluded that to find out how individuals get affected by these mutations and what factors are involved in causing such mutation, a large scale study should be conducted.In addition, other genes involved in causing epilepsy should also be investigated in local Pakistani Punjab population. As a result of such studies, various diagnostic procedures, strategies for counseling and gene therapies can develop.
Availability: Items available for loan: UVAS Library [Call number: 2925-T] (1).
133.
Genetic Evolution And Development Of Recombinant Vaccine Against Newcastle Disease For Chicken In Pakistan
by Abdul Wajid (2009-VA-705) | Dr. Muhammad Wasim | Prof. Dr. Tahir Yaqub | Dr. Muhammad Tayyab.
Material type: ; Format:
print
; Literary form:
not fiction
Publisher: 2017Dissertation note: Newcastle disease (ND) is one of the most contagious diseases of poultry worldwide. The
disease is endemic in Pakistan and recurrent outbreaks have been reported in commercial poultry
flocks, domestic pet and migratory birds since 1963 an inception of commercial poultry farming
in the country. Disease surveillance is necessary to determine the incidence of the disease as well
as to identify the etiological agent of the disease status in the region. The analysis of the field
data provides a clue for the higher authorities to take steps for the remedy of the devastating
outbreak. A virulent form of Newcastle disease virus caused an outbreak in the northern region
of Pakistan during the mid of 2011. The virus was identified as a virulent viscerotropic vvNDV
and characterize, belonging to the sub genotype VIIi. However, the virus of this genotype is still
circulating in the field though the intensity of the strain to succumb the chickens to cause
mortality does not exist. The particular thing in this genotype was its susceptibility to other avian
species like pheasants, peafowls, ducks turkeys, peacocks, sparrows and parakeets. As this
genotype is circulating since 2011 2016 and still spill over in these avian species. Thus for the
last five years (2011-16), 3500 healthy, diseased and dead chickens, pheasants, peacocks,
turkeys, peafowls, ducks, sparrows, exotic parakeets, rosy-faced parrots, pigeons, and partridges
from 750 different locations s were monitored. Samples were collected from the Northern region
of the country Punjab, Khyber Pakhtoonkhawa, Azad Kashmir, including Gilgit,Baltitssan and
from Southern region, Karachi, Hyderabad , Mirpursakro and other small cities where the poultry
farms are located. The samples were collected by the local veterinarians, poultry Assistants and
Animal health practitioners who assist during the surveillance program. Samples were also
collected from the farmers who brought their birds for inspection in the lab with the details of the
141
farm. Mostly sampling was done where there was reports of NDV outbreak, tissues were
collected usually the trachea, spleen and brain, moreover, the pharyngeal and cloacal swabs not
only from the infected birds but also from the healthy birds were collected to assess the virus
shedding in the flock. Blood samples were also collected (1% of the birds at farm), for serum
collection to assess the immune status of the flock using Haemagglutination Inhibition (HI) test
and Enzyme linked immunosorbant assay (ELISA). The Survey Form meet the international
standard was filled for each farm for recording the information required to find the diagnostic
clue as well as the molecular characterization of the isolates. Pool of five pharyngeal swabs were
processed after the passage into 9-day old chicken embryonated eggs and confirming the positive
HA test and then confirmed by real time PCR (RT-PCR). In addition, sera were tested against
NDV by HI and ELISA tests. The targeted samples were sequenced by complete fusion gene and
whole genome using 22 pairs of overlapping primers. The observations indicated that the
commercial broiler industry is highly susceptible to virulent NDV and confirmed by data
available in the laboratory in the survey form. Contrary to that a little is known regarding the
maintenance and enzootic trends of vNDV infection level in domestic and wild birds. Poor
strategy of the use of vaccines and vaccination as well as the existence of virulent form of NDV
in the domestic and pet birds indicate a possibility of the root cause of the ND eruption in the
developing countries. A continuous isolation of virulent viruses of the panzootic Newcastle
disease virus of sub-genotype VIIi since (2011-2016 from commercial chickens and from various
other avian species in the country provide evidence for the existence of epidemiological links
intermingling of the strain among them. Therefore, to avoid the huge economical losses in the
commercial poultry the second largest industry in Pakistan, their close proximity should be
strictly avoided. The mass vaccination of the poultry flocks is in practice in all commercial
142
poultry farms in Pakistan. However, the use and availability of a reliable and standard vaccine, as
well as the correct usage of vaccine dose of the live attenuated LaSota vaccine are the key factors
to improve their efficacy in the field. Minor outbreaks have been occurring in the field even
though a severe outbreak was occurred in 2011-12 collapsed the poultry industry with other pet
and wild birds. To minimize the continuity of these minor outbreaks in the field for long time
there is a need for more effective vaccine to control the particular genotype of the ND virus. In
the present study, DNA vaccine was developed using the SFR-55 NDV strain antigens, in the
form of fusion (F) and hemagglutinin-neuroaminidase (HN), namely pcDNA3.1-F and
pcDNA3.1-HN. In vitro expression of both genes construct was assessed by reversetranscriptase-
PCR (RT-PCR) and western blotting. In the trial an inactivated oil-based emulsion
vaccine was prepared using the field strain SFR-55 and compare with the commercial vaccine
LaSota strain commonly used by the poultry industry. Birds were divided into six groups, the
first two groups were immunized with pcDNA3.1-F and pcDNA3.1-HN alone respectively and
third group with was vaccinated with both antigens pcDNA3.1-F+HN. The other two groups
were immunized with inactivated (wvSFR-55) and LaSota vaccines as described above, the last
group was injected with empty vector as control. The birds were immunized twice at 14 and 21
days of age intramuscularly (DNA vaccine), subcutaneous and eye-drop by inactivated and
LaSota vaccines respectively. The birds were challenged with live virulent NDV strain using a
dose of 10,000 ELD50/0.1ml per chicken. Results indicate that Inactivated and LaSota vaccines
provided high protection (>80%), as compared to pcDNA3.1-F, pcDNA3.1-HN, pcDNA3.1-
F+HN gave 70%, 75% and 20% respectively. There was 100% mortality in control chickens.
The administration of two vectors expressing F and HN antigens induced high immune response,
and provide protection than when used separately. However, the groups immunized with
143
pcDNA3.1-F, pcDNA3.1-F+HN and inactivated vaccine resulted in lower amount of virulent
virus shed after challenge when compared to the group immunized with standard LaSota. In
summary, the co-administration of both NDV glycoprotein antigens increased protection than
use separately. DNA-based vaccine can be used safely to reduce mortality and most importantly
lower the risk of virus transmission due to decreased level of virulent virus shedding. Availability: Items available for loan: UVAS Library [Call number: 2910-T] (1).
134.
Identification Of Pkhd1 Gene Mutation In Polycystic Kidney Disease And In-Silico Molecular Characterization In Different Mammals
by Taslim Un Nisa (2015-VA-1105) | Dr. Wasim Shehzad | Dr Muhammad Yasir Zahoor | Prof.Dr. Aftab Ahmad Anjum.
Material type: ; Literary form:
not fiction
Publisher: 2017Dissertation note: Fibrocystin is a large, receptor-like protein that is involved in the tubulogenisis and maintenance of duct-lumen architecture of epithelium. Fibrocystin has a combination with the primary cilia of epithelial cell. Renal tubules (small tube) of kidney where urine is formed lined by tiny hair like projection.
Twenty five suspected patient was selected and DNA extracted through organic extraction method from the suspected patient blood. Primers were designed PKHD1gene’s coding sequence located at 6p12.2 in human. The coding region sequenced using the ready mate terminator Sequencing Reaction Kit by Perkin Elmer/ABI and read in an automated sequencer. The allele’s variants have been only reported for Fibrocystin protein in human.
All of the sequences are evaluated by using Chromas and Bioedit software for sequence analysis. The in-silico protein analysis is done for normal and mutated alleles through UCSC and RAPTROX. Homology analysis was also done between human and mammals DNA sequence.
We found mutations which are associated with ARPKD disease and these variants are most common in other population whereas we also found some new variants. There are some reported mutations which we found in our study such as (c3790C>T),(c3891G>T),(c3790C>T). We found three new mutations in PKHD1 gene. The new mutations which we found are (c3681G>A),(c3804C>T),(c3931A>C).These mutations (c3790C>T) and (c3931A>C) in the exon 32 show significant effect on the gene and protein function. Geneticanalysis of PKHD1gene show thatPakistanifamilies have mutations as compared to other population along with some common exonic regions such as exon 32 whichisalsodescribed by others in two different studies.We also analyze the pedigrees of these patients which are consanguineous families and autosomal recessive polycystic disease.
We found total six mutations in this gene including missense/ synonymous mutations. In which, three novel mutations and others are reported mutations. These variations from the results are due to the population and consanguineous families’ pattern.In our study, we also found that Mouse and Chickencan preferably be used as a modal organisms in pathology.This study will also help us in the development of molecular genetic testing for their detection in Pakistani families and population.
Availability: Items available for loan: UVAS Library [Call number: 2941-T] (1).
135.
Association Of Tryptophan Hydroxylase 2 Gene Polymorphisms With Risk Of Depression In Homo Sapiens And Equus Caballus
by Sher Sarmad (2015-VA-1062) | Dr. Asif Nadeem | Dr. Maryam Javed | Prof. Dr. TahirYaqub.
Material type: ; Literary form:
not fiction
Publisher: 2017Dissertation note: In the biosynthetic pathway for brain serotonin, Tryptophan hydroxylase-2 (TPH2) acts as the rate-limiting enzyme. It is a key element in maintaining adequate serotonin neurotransmission in the central neuron system (CNS). It is broadly discussed as an important candidate gene in multiple psychiatric disorders, especially suicidal behavior and depression. A relationship between TPH2 and major depressive disorder (MDD) has been reported by multiple gene-disease association studies in different populations. Horse can be employed as a valuable candidate for an animal model of depression because it shares environmental factors which are known to cause depression in humans.The hypothesis of this study was that, there is association between TPH2 gene polymorphisms and risk of developing MDD.Blood was collected from each participant.Human (Experimental and Control group) and Horse (Experimental and Control group).DNA wasextracted usingthe standard Phenol Chloroform Isoamyl alcohol (PCI) protocol. Specific set of primers were designed for the amplification of TPH2 gene exons and partial region of the introns. The amplified PCR products was precipitated and sequenced for the identification of variants. For sequence data analysis Chromas Software(2.1) was used along with BLAST available at NCBI and Clustal W program. Multiple alignments were performed for polymorphism identification and association of identified polymorphisms was performed using SPSS.The significant association of the variant rs7305155 with Major Depressive Disorder indicates that TPH2 has a role in disease etiology.Understanding the extent of the role different genes play in MDDmay help in tailoring medication. “Gene-Response to drug” association studies have also shown that patients with certain polymorphisms might respond better to certain class of drugs. It is useful to know which variants are prevalent in our population. Availability: Items available for loan: UVAS Library [Call number: 2939-T] (1).
136.
Analysis Of Tp53 Gene Isolated From Oral Cancer Patients
by Amir Saleem (2013-VA-897) | Dr. Muhammad Wasim | Prof. Dr. Tahir Yaqub | Prof. Dr. Habib-ur-Rehman.
Material type: ; Literary form:
not fiction
Publisher: 2017Dissertation note: Cancer is a term used for disease in which abnormal cells divide without control and are able to invade other tissues. Cancer cells can spread to other parts of the body through blood and lymph system.
TP53 is one of the most important tumor suppressor genes, mutated in more than 50% of human malignancies. It controls DNA repair, cell cycle and apoptosis and therefore plays an essential role in keeping genetic constancy. TP53 gene is present on the short arm of chromosome number 17. In human it extends 19,200 bp in 11 exons.
Various parameters are used in the present study was aimed to investigate coding regions of TP53 gene for analyzing the mutations involved in oral cancer. Human OSCC samples (15) and normal tissue samples (15) were collected from Fatima Memorial Hospital Lahore. Samples were collected in tubes by oncologist containing ethanol and then brought to Molecular Biology and Biotechnology Lab of Institute of Biochemistry and Biotechnology, UVAS Lahore and frozen at -20°C before DNA extraction. Samples have been processed for research purpose. DNA was extracted from tissue by using Trizol Method and quantity was checked by nanodrop spectrophotometer. Two Primer sets were designed to amplify protein coding region of TP53 gene. After amplification through PCR, DNA Sequencing was done. Data interpretation was done by using several softwares like BLAST alignment tool, Bioedit, Clustal W2.
In this study it was tried to find out the mutations in TP53 but no any kind of mutations were identified. Because I conducted my research only on 15 samples. So in future to use this gene as a potentional biomarker we can increase our number of samples.
46
The need for today is to develop valid biomarkers, which can be incorporated in ongoing in vivo and in vitro clinical mechanistic and improve the diagnosis and prognosis of this dreadful disease.
In the conclusion, we must say that further study, with much larger sample number, is required to revise the effects of this polymorphism and accurately identifying the associated factors. There is a need to explore the other gene mutations causing oral cancer in local population of Punjab and Pakistan that will ultimately help to develop genetic counseling strategies, gene therapies and prenatal diagnostic procedures for the population of Pakistan. Availability: Items available for loan: UVAS Library [Call number: 2937-T] (1).
137.
Polymorphism Analysis Of Bovine Tumor Necrosis Factor Alpha Gene And Its Association With Mastitis In Nili Ravi Buffaloes
by Samia Tanveer (2011-VA-362) | Dr. Sehrish Firyal | Dr. Muhammad Wasim | Dr. Muhammad Nawaz.
Material type: ; Literary form:
not fiction
Publisher: 2017Dissertation note: Various number of factors cause hindrance in the milk production potential of buffalos. Mastitis is the costly and most prevalent disease causing production losses of dairy herds in Pakistan and elsewhere in the world. Susceptibility and resistance to mastitis is complex trait influenced by genetic variation of animals. Among these immunity gene variations, the polymorphism in tumor necrosis factor alpha gene (TNF-α) play important role in immune response to virus. Polymorphism in TNF-α gene is associated with mastitis susceptibility and resistance. It would be a potential candidate gene for imparting resistance mastitis in dairy buffalos. Blood sample were taken from the 20 Nili Ravi buffalos having clinical and subclinical mastitis. Extraction of DNA was done from frozen blood after thawing, using organic extraction method & also kit method followed by DNA quantification (i.e. gel electrophoresis and nanodrop). Total 5 primers were designed using Primer3 bioinformatics tool. All these primers were optimized using different protocols and a set recipe was obtained for each primer. The amplification of DNA samples was done one by one using all these five primers on optimized protocol. The amplicons obtained were subjected to agarose gel electrophoresis to check whether we have the required product or not using 100 kb ladder and then amplicones were send for the sequencing.
Summary
110
The sequencing analysis of resulted amplicon sequence was done using Bioinformatics software Finch TV. Total of 6 mutations were found while 5 were same in all the samples whereas 6th mutation was found only in clinical samples. It is valuable in accomplishing genetic progress for resistance and to improve the immune response. This study will paved the way for animal breeder for selection of Nili Ravi mastitis resistant buffalos for breeding. TNF-α gene polymorphism based marker is now available for screening of resistant bulls as well. Availability: Items available for loan: UVAS Library [Call number: 2936-T] (1).
138.
Bacterial Profiling And Development Of Molecular Diagnostic Assays For Detection Of Bacterial Pathogens Associated With Bovine Mastitis
by Aqeela Ashraf (2012-VA-388) | Dr. Muhammad Imran | Prof. Dr. Tahir Yaqub | Dr. Muhammad Tayyab.
Material type: ; Literary form:
not fiction
Publisher: 2017Dissertation note: The livestock sector plays a critical role in strengthening the economy of Pakistan. Control of livestock diseases is the primary objective of government livestock departments. Bovine mastitis is among the most significant diseases of livestock as reported by various field surveys in Pakistan. Despite considerable knowledge about mastitis and its etiology, this disease is still prevalent in many dairy herds; it remains most difficult to eradicate or control. It is an inflammation of mammary gland resulting in decreased milk production, veterinary care costs and culling losses.
In animal health improvement, there is a paradigm shift from treatment of clinical illness to disease prevention. Recognition of disease is the foundation of disease control and prevention. California mastitis test and somatic cell counting are the most commonly used methods for diagnosis of bovine mastitis. These methods are unable to identify the causative agent. Detection of pathogen is critically important for better control of mastitis. Microbial culturing and biochemical tests are traditionally used methods for pathogen identification. But, these methods are very time consuming and can only detect viable bacteria from the sample and can lead to false negative results. The progress in molecular methods based on PCR has improved the veterinary diagnostics.
For the identification of bovine mastitis pathogens, an economical, rapid and sensitive molecular diagnostic assay was developed using multiplex PCR, detecting the pathogenic species-specific DNA. The target species areS. aureus,E. coli, S. uberis, S. agalactiae, S. dysagalactia, S. haemolyticus, S. epidermidis, S. chromogenes andM. bovis. Multiplex PCR assay was developed for the detection of these significantly important bacterial pathogen
causing bovine mastitis. Species specific primers were designed which have the ability to specifically amplify the particular gene in the target species. For this purpose various gene regions were selected for different bacterial species which included 16S rRNA, cpn60, phoA and rdr. Initially monoplex PCRs were optimized individually for each target species. For optimizing multiplex PCR assay, various combinations of individual PCRs with varying concentrations of primers and template DNA were used. The final protocol included all the nine sets of primer pairs, every set targeting a unique mastitic pathogen. Multiplex PCR assay was checked for its specificity and analytic sensitivity was calculated. Mastitic milk samples were collected aseptically from various farms. Initial screening was based on Surf field mastitis test and California mastitis test. Milk samples were cultured on nutrient agar, blood sheep agar and MacConkey’s agar. The bacterial isolates were identified and further sub-cultured in nutrient broth. All the isolates were identified on the basis of 16S rRNA sequencing analysis.
The developed multiplex PCR assay was used to detect the target bacterial pathogens from the collected milk samples. Limit of detection of developed assay was up to 50 pg for DNA isolated from pure cultures and 104 CFU/ml for spiked milk samples. The results obtained by 16S rRNA sequencing, bacterial culture based identification and multiplex PCR assay were compared. Sensitivity and specificity were calculated using latent class analysis, specificity was up to 88% and sensitivity was up to 98% for targeted mastitic pathogens. The developed multiplex PCR detected nine bacterial species in a single reaction. Multiplex PCR assay has also detected the bacterial pathogens in a few culture-negative mastitis milk samples. This is the first multiplex PCR assay which can efficiently detect nine important mastitic bacterial pathogens in a single reaction. The development of multiplex PCR assay is useful in early diagnosis and prevention & control of bovine mastitis.
Mycoplasma is often ignored as a major mastitis-causing pathogen due to lack of rapid and accurate diagnostic tools. In this study a LAMP assay was developed for the identification of M. bovis from clinical mastitic milk samples. LAMP primers were designed from three gene regions including uvrC, 16S rRNA and gyrB. Bacterial reference strains and mastitic milk samples positive for M. bovis were collected from Quality Milk Production Services, Cornell University, Ithaca, NY. Bacterial strains were further cultured on Hayflick medium containing 15% horse serum and incubated for up to 7 d at 37°C with CO2 enrichment. DNA was isolated from mastitic milk samples and bacterial culture using Qiagen DNeasy Blood and Tissue Kit (Life Technologies, Carlsbad, CA) according to the manufacturer’s instructions with few modifications. PCR and LAMP assay was performed for all the samples obtained. Analytic sensitivity was calculated and the limit of detection was up to 50pg/reaction for LAMP assay which is higher as compared to PCR.
Sensitivity and specificity was calculated for each of the three tests. Cohen’s kappa values obtained were 0.940 for uvrC, 0.970 for gyrB and 0.807 for 16S rRNA. All three tests showed a high level of agreement between test results and the true mastitis status, indicated by the receiver operating characteristic (ROC) curve. A robust, sensitive and specific LAMP assay has been developed for the detection of M. bovis from mastitic milk. These novel molecular assays could be helpful for correct and timely identification of bovine mastitic pathogens, which is crucial for the control and treatment of the disease.Molecular diagnostic assayshave been developed in the current study based on multiplex PCR assay and loop-mediated isothermal amplification assay.
Availability: Items available for loan: UVAS Library [Call number: 2930-T] (1).
139.
Molecular cell biology
by Darnell, James E | Lodish, Harvey F | Baltimore, David | Berk, Arnold.
Edition: 8th ed.Material type: ; Literary form:
not fiction
Publisher: USA: Macmillan Learning; 2016Availability: Items available for loan: UVAS Library [Call number: 571.6 Darnell 32408 8th 2016 Genetics] (1).
140.
Genetic Identification And Molecular Classification Of Birds Commonly Found In Lahore Through Dna Barcoding
by Agha Wasif Ali Khan(2014-VA-01) | Dr.Ali Raza Awan | Dr.Muhammad Wasim.
Material type: ; Format:
print
; Literary form:
not fiction
Publisher: 2017Dissertation note: T Availability: Items available for loan: UVAS Library [Call number: 2938-T] (1).
141.
Human molecular genetics
by Strachan, T | Read, Andrew P.
Edition: 4th ed.Material type: ; Literary form:
not fiction
Publisher: USA: Garland Science; 2011Availability: Items available for loan: UVAS Library [Call number: 611.01816 Strachan 32456 4th 2011 Genetics] (1).
142.
Cell and Molecular Biology
by Vyas, S. P | Mehta, A.
Edition: 1st revised ed.Material type: ; Literary form:
not fiction
Publisher: India: CBS Publishers & Distributors; 2014Availability: Items available for loan: UVAS Library [Call number: 571.6 Vyas 32476 1st 2014 Genetics] (1).
143.
Molecular Histochemical Techniques
by Koji Takehiko.
Edition: 1st ed.Material type: ; Literary form:
not fiction
Publisher: Germany: springer; 2000Availability: Items available for loan: UVAS Library [Call number: 572.8 koji 32522 2000 Molecular histochemical] (1).
144.
Introduction to Genetic Analysis
by Griffiths, Anthony J.F | Wessler, Susan R.
Edition: 10th ed.Material type: ; Literary form:
not fiction
Publisher: USA : W. H. Freeman, 2012Availability: Items available for loan: Pattoki Library [Call number: 576.5 Griffiths 50522 11th 2015 Genetics] (2), UVAS Library [Call number: 576.5 Griffiths 32608 10th 2012 Genetics] (1). Checked out (1).
145.
Plant biochemistry /
by Gleason, Florence K | Chollet, Raymond.
Material type: ; Format:
print
; Literary form:
not fiction
Publisher: Sudbury, Mass. : Jones & Bartlett Learning, c2012Availability: Items available for loan: Pattoki Library [Call number: 572.2 Gleason 50005 1st 2012 Botany] (1).
146.
Molecular Biology and Biotechnology
by Rasad, Dr. D. Vijay Raghava.
Edition: 1st ed. Material type: ; Literary form:
not fiction
Publisher: New Delhi: ASTHA Publishers & Distributors; 2015Availability: Items available for loan: UVAS Library [Call number: 572.8 Prasad 32680 1st 2015 Biotechnology] (1).
147.
Principles and techniques of Biochemistry and molecular Biology
by Hofmann, Andreas | Clokie, Samuel.
Edition: 8thMaterial type: ; Literary form:
not fiction
Publisher: New Delhi: Cambridge ; 2018Availability: Items available for loan: UVAS Library [Call number: 573.44 Hofmann 33039 8th 2018 Biochemistry] (2).
148.
Molecular Cell Biology / 8th ed
by Lodish, Harvey | Zipursky, S. Lawrence | Matsudaira, Paul | David, Baltimore | Darnell, James | Lodish, Harvey F | Berk, Arnold.
Edition: 8th ed.Material type: ; Literary form:
not fiction
Publisher: U.S.A: W H Freeman & Co; 2016Availability: Items available for loan: UVAS Library [Call number: 571.6 Lodish 33117 8th 2016 Genetics] (1).
149.
Encyclopedia of molecular cell biology and molecular medicine
by Meyers, Robert A.
Edition: 1st ed.Material type: ; Literary form:
not fiction
Publisher: USA: Wiley; 2004Availability: Items available for loan: UVAS Library [Call number: 572.803 Meyers 18243 Vol.4 2004 MolecularBiology] (5).
150.
Encyclopedic reference of genomics and proteomics in molecular medicine
by Ganten, D | Ruckpaul, Klaus.
Edition: 1st ed.Material type: ; Literary form:
not fiction
Publisher: Germany: Springer; 2006Availability: Items available for loan: UVAS Library [Call number: 616.04203 Ganten 20089 Vol.1 2006 MolecularBiology] (1).
