African Journal of Biotechnology Research

African Journal of Biotechnology Research ISSN 9316-2194 Vol. 4 (10), pp. 221-228, October, 2016. © International Scholars Journals 

Full Length Research Paper

Sequence SSR marker DNA clones G. mangostana of ISSR-suppression-PCR technique

Joko L. Malik*1, Uwais K. Suhurto2, Adams Raffi Atatas1, Ali Ahmed Habibie3

1Laboratory of Plant Breeding, Faculty of Agriculture, University of Padjadjaran, Indonesia

2Center for Agriculture Biotechnology, Kasetsart University, Thailand

3Gene Research Center, University of Tsukuba, Japan.

Email: joko.maliq3@hotmail.com

Accepted 19 September, 2016

Abtsract

Assessment and utilization of diversity in plant genetic resources is very important to improve of plant species. A mangosteen (Garcinia mangostana L.) seed was formed obligate apomicts process. Genetic diversity of mangosteen was developed by using SSR markers. An inter-simple sequence repeat (ISSR)-suppression - PCR technique established to SSR marker of plant spesies. DNA library construction was used restriction enzyme blunt end Rsa I and adaptor (consist of 48-mer: 5’-GTAATACGACTCACTATAGG GCACGCGTGGTCGACGGCCCGGGCTGGT-3’ and 8-mer with the 3-end capped by an amino residue: 5’-ACCAGCCC-NH2-3). Primer designed by using compound SSR (AC)10; (TC)6(AC)5 or (AC)6(AG)5 and an adaptor primer AP2 and nested PCR using AP1 primers. The PCR product integrated into the plasmid PGEMT Easy Vector System and competence cell Escherichia coli (strain DH5α) and sequence. Eight sequence from sample #G17 with SSR compound (AC)10, (TC)6(AG)5 and (AC)6(AG)5 produced two primer pairs. The primer pairs from sequence positive clone 4 of SSR compound (TC)6(AC)5 were forward primer: 5’-GGCCGTT AAAGTAGCTCAAGAA-3’ and reverse primer:5’-CCGCATAGCATCAGTATCTG TC-3’, while primer pairs from sequence positive clone 10 of SSR compound (AC)6(AG)5 were forward primer: 5’-GTGTTTCCATTTGTTACGCGCT-3’ and reverse primer: 5’TAATGCCGTTGGGCAGTGA-3’ 

Keywords: Garcinia mangostana, SSR compound. SSR primer