Livestock is playing a major role in the uplift of our national economy in terms of revenue from milk, meat and hides. In spite of this major contribution this sector is facing hurdles in its development. The infectious diseases and their vectors have occupied a mainstay in posing the uplift of this sector. Boophilus is an important biological vector for various protozoan and bacterial infections in animal as well as human. To date the identification of these ticks mainly rely on the morphological basis which have many variations among different identification keys. To make the identification more accurate at species level, the use of molecular tools is very necessary. Ticks were collected from the various areas of district Peshawar through random convenient sampling method. Briefly, 50 cattle and 50 buffaloes were sampled through forceps. Various ticks spp. were stored in 70% ethanol for further processing. Among the species Rhepicephalus microplus (Boophilus microplus) was the most prevalent specie (25.64103%) followed by Rhepicephalus annulatus (5.413105%) Rhipicephalus decloratus (5.128205%) Rhipicephalus distinctus (4.273504%), Rhipicephalus arnoldi (3.988604%), Rhipicephalus evertsi (5.698006%), while in Heamaphysalis species Heamaphysalis aciculifer highly prevalent (5.128205%) followed by Haemaphysalis parmata (4.843305%), Haemaphysalis excavatum (3.988604%) and Haemaphysalis houyi (3.988604%), so far Hyalomma species is concerned includes Hyalomma anatolicum (3.988604%), Hyalomma trancatum (4.843305%), Hyalomma detritium (5.982906%), Hyalomma egyptium (4.273504%), Hyalomma impeltatum (0.854701%) Hyalomma rufipes (1.709402%), Amblyomma pomposum (4.273504%), Dermacentor rhinocerinus (2.849003%), D. circumguttatus (3.703704%), and
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Dermacentor marginatus (2.564103%) are highly prevalent in cattle. Among the buffalo, Rhipicephalus 173 (43.25 %) followed by Haemaphysalis 82 (20.5 %), Hyalomma 54 (13.5 %), Dermacentor 26 (6.5 %) and Amblyomma 9 (2.25 %). The species prevalent in Rhipicephalus are Rhipicephalus microplus 74 (42.78%), Rhipicephalus annulatus 15 (8.68%), Rhipicephalus decloratus 19 (10.99%), Rhipicephalus distinctus 14 (8.10%), Rhipicephalus arnoldi 16 (9.25%), Rhipicephalus evertsi 17 (9.84%) and Rhipicephalus kochi 18 (10.40%) followed by Haemaphysalis aciculifer 18 (21.96%), Haemaphysalis parmata 15 (18.30%), Haemaphysalis excavatum 23 (28.05%), and Haemaphysalis houyi 26 (31.70%), so far Hyalomma species is concerned, Hyalomma anatolicum 10 (18.52%), Hyalomma tranctum 7 (12.96%), Hyalomma detritium 9 (16.67%), Hyalomma egyptium 7 (12.96%), Hyalomma impeltatum 10 (18.52%), and Hyalomma rufipes 11 (20.37%) and Dermacentor rhinocerinus 9 (34.62%), followed by Dermacentor circumgutattus 8 (30.76%) and Dermacentor marginatus 9 (34.62%) and Amblyomma is concerned Amblyomma pomposum 9 (2.25%). DNA was extracted from the Rhipicephalus (Boophilus) ticks through phenol-chloroform method. The extracted product was then run by gel stained ethidium bromide. The gel was visualized and examined bands on UV illuminator. Different sequences were retrived from database and genus specific primer were designed for the amplification of ITS-2 gene of Rhipicephalus genus of hard ticks. A consensus sequence was retrieved, a set of primers were designed by using Bioedit softwere version 7.2.6. DNA was extracted from 100 ticks and then run by PCR. Specific primers were designed for ITS2 gene. Phylogenetic tree based on the DNA sequences amplified from extracted from all the comparison with ticks and determined Genus Rhipicephalus area that are ITS2 Rhipicephalus ITS2 ribosomal RNA gene sequence 18s, thus obtained from Genebank.