ISSN-2319-2119 RESEARCH ARTICLE Ernieenor Faraliana et al, The Experiment, 2013 Vol. 15(2), 1064-1071 www.experimentjournal.com 1064 PCR AMPLIFICATION OF MITOCHONDRIAL CYTOCHROME B GENE OF ANIMALS IN MALAYSIA ABSTRACT The aim of this study is to acquiring basic data for identifying blood meal species of vector arthropods in Malaysia by amplify and sequence the mtDNA cytb gene of captive and domestic animals. Polymerase Chain Reaction (PCR) using universal primers complementary to the conserved region of the mitochondrial DNA (mtDNA) cytochrome b (cytb) gene fragment, was performed on DNA of blood samples of 27 captive and domestic animals in Malaysia. DNA of hosts was amplified by PCR and the products were visualized on gel electrophoresis. Twenty two sequences were obtained and compared with sequences registered in GenBank using a BLAST program. The percentage of similarity between the study species and GenBank species was in the range of 78 - 100%. Four sequences had no significant similarity and 3 species were mismatched with other species in the range of 78 - 89% similarity. Nine new gene sequences with accession number JQ812112 (Prionailurus bengalensis), JQ812113 (Prionailurus bengalensis), JQ812114 (Tapirus indicus), JQ812115 (Oryx dammah), JQ812116 (Macaca nemestrina), JQ409474 (Panthera tigris 2), JQ409475 (Panthera tigris 1), JQ409476 (Panthera tigris 4) and JQ409477 (Malayan Elephant) had been deposited in GenBank. These results will contribute to develop the GenBank database and can be applied for further epidemiological studies on vector borne diseases in tropical countries including Malaysia. keywords Cytochrome b gene, Polymerase Chain Reaction, GenBank, Mitochondrial DNA, Arthropods. 1. INTRODUCTION Majority of pathogens survive in nature by utilizing animals as their vertebrate hosts. The organisms are taken up by arthropod vectors from infected hosts and transmitted either to an intermediary host or directly to a susceptible human host [1]. Some of vector-borne diseases like West Nile virus use an intermediary animal host to serve as a reservoir for the pathogens until susceptible human populations are exposed to the disease. Increased contact between man and wild animals due to environmental changes and deforestation has increased the risk for contracting certain arthropod-borne diseases [2]. Captive animals in zoos and domestic animals may serve as feeding hosts for arthropods. These animals may be harboring pathogens of public health importance. It is important to identify natural hosts of the pathogens as fast and accurate possible for an effective outbreak management and control. If the vertebrate host of a certain disease is known, control measures can be implemented and transmission of disease prevented as early as possible. In the past, identification of the blood meals of haematophagous arthropods by serological techniques such as the precipitin test, latex agglutination test and enzyme-linked immunosorbent assays (ELISA) were commonly used for a wide variety of purposes [3, 4, 5, 6, 7]. While those techniques continue to provide valuable and insightful data, the identification of many arthropod blood meal sources is limited to only order or family level [8]. A technique is therefore needed to further identify the blood meal sources beyond this level. The availability of DNA sequence data of various vertebrates has opened the door for molecular-based blood meal analysis approaches, such as Polymerase Chain Reaction (PCR) [9, 10, 11]. The alternative method of PCR-based identification of vertebrate host blood meals is more convenient and easier to perform than the previously used serological tests. The most straightforward and specific method to identify blood meals is sequencing of amplified DNA. This approach is ideal since primers can be employed to amplify conserved homologous DNA fragments from diverse potential of vertebrate blood sources [8]. Nowadays, mitochondrial DNA (mtDNA) cytochrome b gene (cytb) is widely used in identifying the species of an organism. The mtDNA contains high proportion of evolutionary-caused nucleotide substitution making it particularly valuable in discriminating