a SciTechnol journal Research Article International Journal of Evolution Singla et al., Int J Evol 2013, 2:1 http://dx.doi.org/10.4172/2324-8548.1000101 All articles published in International Journal of Evolution are the property of SciTechnol, and is protected by copyright laws. Copyright © 2013, SciTechnol, All Rights Reserved. International Publisher of Science, Technology and Medicine Genetic Relationship Among Indian Termites Based on DNA Sequence of Mitochondrial 12S Ribosomal RNA Gene Mandakini Singla 1 , Vijay Lakshmi Sharma 2 , Ranbir Chander Sobti 3 *, Monika Sodhi 4 and Mamtesh Kumari 1 Abstract Partial 12S gene fragments were amplifed by using specifc primers in nine species of termites of the genus Odontotermes, Microtermes and Microcerotermes (Isoptera: Termitidae: Macrotermitinae), and the PCR products were subjected to sequence analysis. The sequences obtained were characterized to see the frequencies of each nucleotide bases, and high A+T content was observed. The divergence of the species was found be lower within the same family, and highest with species from the family Rhinotermitidae. Phylogenetic tree drawn on the basis of distance Neighbour-joining method revealed clustering of individuals according to their genera and families. Keywords Macrotermitinae; Phylogeography; Termites; 12SrRNA Introduction Insect molecular systematics has complemented and enhanced the value of morphological and ecological data, making substantial contributions to evolutionary biology in the process. During the past decade, our understanding of the relationship among organisms at various levels of taxonomy has advanced greatly with the aid of DNA molecular systematic techniques and phylogenetic theory. However, the advent of molecular taxonomy provides more sensitive techniques for examining complex identifcation issues. Advances in method objects have led to the accumulation of large amounts of DNA sequence data from most major insect groups [1]. Molecular taxonomy based on mitochondrial DNA has proved to be an efcient alternative to species identifcation and their phylogenetic relationships. Infact, mitochondrial markers have been used with a number of insects for systematic and identifcation purposes [2,3]. Te use of mitochondrial genome sequence is further supported by the occurrence of cladistically informative gene order rearrangement events. Mitochondrial sequence data have, therefore, been extensively used in the past 10 years to evaluate the population structure, gene fow, phylogeny, phylogeography and taxonomy of termites [4]. Indeed, several studies based on mitochondrial genome *Corresponding author: Ranbir Chander Sobti, Department of Biotechnology, Panjab University, Chandigarh, India, Tel: 91-172271078; E-mail: rcsobti@pu.ac.in Received: December 07, 2012 Accepted: February 20, 2013 Published: February 23, 2013 sequences such as the cytochrome oxidase genes and the AT rich region have thrown a great deal of light on termite taxonomy. Termites are also very important ecological players in tropical ecosystem, having been described as “ecosystem engineers”, due to their important role in providing soil ecosystem services [5]. Termites adapt to arid environment and play an important role in decomposition, where common decomposers such as micro bacteria and fungi cannot function. Despite their importance, our understanding of a number of their basic biological processes in termites is extremely limited. Developing a better understanding of termite biology is closely dependent upon reliable species identifcation. Te use of molecular markers may be helpful in estimating phylogenetic relatedness between the termite species and estimating genetic diferentiation among local populations within each species. 12SrDNA is highly conserved, and has been employed to illustrate phylogeny of higher categorical levels such as in phyla or subphyla. In general, 12S and 16S rDNAs are the most conserved regions among the mitochondrial genes. Mitochondrial 12S and 16SrRNA genes are useful for the phylogenetic studies because of their slow evolutionary rate, and the existence of universal insect primers and ease of reliable PCR amplifcation. In the present study, the molecular characterization of nine species of termites of family Termitidae has been described, because very scarce information is available in relation to the mitochondrial 12SrRNA in termite species. Te data on this gene in Indian termites is also not available. Materials and Methods Collection and storage of samples Te specimens of the termites under study were collected from various locations in India (Table 1). Te voucher specimens were preserved in absolute ethanol mixed with a few drops of glycerol, and maintained in the Department of Zoology, Panjab University, Chandigarh (UT), India, till the extraction of genomic DNA. Identifcation of termites Soldier specimens of all the species collected (packed in scintillation vials in rectifed alcohol) were got identifed from Zoological Survey of India, Kolkata and Forest Research Institute, Dehradun, using the keys or descriptions of Roonwal and Chhotani [6]. Te details of collection site, date of collection, source and name of the collector were mentioned on each vial. Isolation of genomic DNA (gDNA) gDNA was extracted from worker termites using the modifed phenol: chloroform extraction method [7]. Te whole insect was homogenized in 1.5 ml eppendorf tube in 500 µl of TE (Tris EDTA pH 8), with hand pestle, and the homogenate was centrifuged at 10,000 rpm for 10 min in cooling centrifuge (-4°C). Te supernatant was discarded, and the pellet was dissolved in 500 µl of lysis bufer (400 µl of TE and 100 µl of 5% SDS), followed by the addition of 6 µl of Proteinase K, and the solution was incubated at 65°C for 1 and a half hour in the incubator. A mixture of 120 µl of phenol: chloroform: isoamyl alcohol (25: 24: 1) was added and the tubes were vortexed