DNA Barcoding for Wild Rice [<i>Oryza rufipogon</i> Griff.] of NBU Campus Based on matK gene and Assessment of Genetic Variation Using DREB and BAD2 Gene Sequences

Abstract

DNA barcoding is a technique for characterizing species based on short DNA sequence of a particular genomic region. The chloroplast maturase gene K (matK) sequence of 1420 bp was used to construct unique DNA barcode for wild rice [ Oryza rufipogon Grirr.] of NBU campus using BOLD system. The partial matK gene sequence (1420 bp) was annotated and submitted into the GenBank of NCBI (Accession no. KM516199). Gene sequences of DREB transcription factor and BAD2 were analyzed to assess the genetic variation between O. rufipogon (NBU) and other species of rice. The DREB gene is responsible for abiotic stress tolerance whereas BAD2 is a gene for fragrant formation in rice. A thick and discrete band of 916 bp was detected on 1% agarose gel in the DREB lane which proved that wild taxon may provide abiotic stress tolerance. The DNA sequence of 916 bp of DREB gene of O. rufipogon of NBU campus was run in BLAST program of www.gramene.org database, it was matched with DREB gene sequence of wild rice O. rufipogon in the database and aligned in chromosome 1, 4 and 9. Both the DREB and BAD2 gene sequences were run in MEGA6 software for study of genetic relationship among the rice species. Phylogram constructed from this analysis showed close clustering association with other wild rice species. The species O. rufipogon of NBU campus was non-fragrant homozygous because it was given PCR product of 357 bp while used four primers in allele specific gene amplification method.