55 AJCS 8(1):55-61 (2014) ISSN:1835-2707 SSR based genetic diversity analysis in a diverse germplasm of groundnut (Arachis hypogaea L.) from Pakistan Sohaib Roomi 1* , Bibi Sabiha 2 , Arshad Iqbal 4 , Muhammad Suleman 3 , Izhar Muhammad 4 , Muhammad Amir Zia 4 , Muhammad Zulfiqar Ahmad 4 , Farooq Rashid 4 , Abdul Ghafoor 4 and Nabila Tabbasam 4 1 Department of Biosciences COMSATS Institute of Information Technology, Islamabad, Pakistan 2 Atta-ur-Rahman School of Applied Biosciences (ASAB), National University of Sciences and Technology, Islamabad, Pakistan 3 Institute of Biotechnology and Microbiology, University of Swat, Khyber Pakhtunkhwa, Pakistan 4 National Institute for Genomics and Advanced Biotechnology, National Agricultural Research Centre, Islamabad, Pakistan * Corresponding author: sohaibbiotch@comsats.edu.pk Abstract The current study was aimed to explore the genetic diversity among seventy Pakistani accessions of Arachis hypogaea. In Pakistan their morphological and biochemical variations have already documented but still so far, molecular variations need to be studied for this valuable crop. For molecular diversity study DNA was extracted from all seventy accessions of Arachis hypogaea. The extracted DNA was primed with thirty SSR primers and amplified through PCR. Fifteen out of thirty primers generated polymorphic bands among the selected accessions. In total, forty different polymorphic loci were determined across the selected accessions. The range of number of polymorphic loci detected was ranged from 2 to 4 for each primer, with an average of 2.6 loci per primer. Polymorphic Index Content (PIC) value was calculated for each marker. The dendrogram was constructed on the basis of allelic data from fifteen SSR markers across the selected accessions. All the accessions were divided into six clusters at 0.67 coefficients of similarity. This study of variations at molecular level of Pakistani groundnut accessions will be helpful for conservation and breeding purposes of groundnut and other legumes. Keywords: Simple Sequence Repeat, Polymorphic Index Content, Groundnut, Loci. Abbreviations: PCR_Polymerase Chain Reaction, SSR_Simple Sequences Repeat, PIC_Polymorphic Index Content. Introduction Groundnut (Arachis hypogaea L.) is grown throughout the world as a source of oil and protein. The Arachis hypogaea is an allotetraploid (AABB, 2n = 4x = 40 chromosomes), resulting from duplication of AA and BB wild type species (Leal-Bertioli et al., 2009). A. hypogaea belong to genus Arachis of Fabaceae. Two main classes of A. hypogaea are hypogaea and fastigiata. A. hypogaea have flowers on the main axis while fastigiata lack flowers on the main axis (Krepovickas and Gregory, 1994). The classification of groundnut only on the basis of their morphological characteristics is not sufficient therefore the assessment of variation at gene level during germplasm collections and pedigree construction is necessary. The use of molecular markers will be helpful for the collection of advanced and novel genotypes of groundnut. DNA based markers provide accurate knowledge at gene level which was not possible with phenotypic markers (Altinkut et al., 2003). Molecular markers have been used for tagging of important traits of groundnut in inter specific introgression populations. Many markers have been identified which were resistant to late leaf spot (Mace et al., 2006). Despite significant physiological, agronomic and morphological variation the peanut exhibits variations at low level by RAPD (Mondal et al., 2005), SSR (Raina et al., 2001) and AFLP (Herselman, 2003). The use of microsatellites to track advantageous traits in plant breeding and as helpful point in gene cloning program explores their importance (Brown et al., 1996). Microsatellites were mostly used in genetic diversity studies, gene flow mating system and paternity studies (Rosseto et al., 1999). Being with highest PIC, due to their high mutation rate, SSRs are routinely used in finger-printing Chen and Du, (2006), molecular mapping (Zhao et al., 2005) phylogenetic and genetic relationship studies (Yang et al., 2005) and marker- assisted breeding (Sun et al., 2006). PCR amplified the SSR loci using primers created from distinctive contiguous nucleotides. Polymorphism is due to the differences in the amount of repeats. Point mutations and polymerase slippage are the main reason of variation in number of repeats (Kruglyak et al., 1998). Microsatellites, (SSRs) have been used to study genetic variation and to construct molecular maps in numerous crops (Lee et al., 2004). ATT and CTT are the most common tri nucleotide sequences in plant genomes (Ferguson et al., 2004). Similarly AT is proved to be the most abundant dinucleotide repeat sequences followed by AG/CT and GT/CA in plant genomes (Cuc et al., 2008). The aim of the current study was to explore the genetic diversity at molecular level among accessions of Arachis hypogea from Pakistan. Morphological and physiological variations have been already documented but molecular variation has been not properly studied for this valuable crops. The analysis of variations at molecular level will be helpful for conservation