Fragrance genotyping of local rice germ plasm by using Allele Specific Amplification (ASA) of the BADH2 gene

• Main Author: Khairun Nisaa' Mohd Nor
• Format: Academic Exercise
• Language: English
• Published: 2011
• Subjects: QH Natural history
• Online Access: https://eprints.ums.edu.my/id/eprint/14748/1/ae0000003135.pdf; https://eprints.ums.edu.my/id/eprint/14748/

The occurrence of fragrance in rice has been associated to the accumulation of 2-Acetyl-1-Pyrroline (2AP). The increased level of 2AP is due to an 8 bp deletion and three Single Nucleotide Polymorphism (SNPs) in exon 7 of the Betaine Aldehyde Dehydrogenase 2 (BADH2) gene which is located on chromosome 8. The two types of mutations cause a premature stop codon that disables the production of the BADH2 enzyme. On the other hand, a fully functional copy of the gene is present in non-fragrant rice varieties. Allele Specific Amplification (ASA) assays can be used for the differentiation between fragrant and non fragrant rice varieties. In this study, a total of 98 local rice samples were used to ascertain its fragrance status. A Thailand rice variety, Thai Hom Mali, was used as a positive control for fragrant rice, while another rice variety, IR36, was used as a negative control for non-fragrance. Four primers namely External Sense Primer (ESP), Internal Fragrant Antisense Primer (IFAP), Internal Non-fragrant Sense Primer (INSP) and External Antisense Primer (EAP) were used in the PCR reaction during ASA of BADH2 gene. The ESP and EAP primers served as internal positive controls by amplifying a region of approximately 580 bp in both fragrant and non fragrant varieties. The IFAP primers amplified a region of 355 bp in non fragrant varieties meanwhile INSP. amplified a region of 257 bp in fragrant varieties. Only 53 samples were amplified out of the 96 rice cultivars. From these 53 samples, 43 were homozygous dominant and were categorized as non fragrant. There were two cultivars that were heterozygous and were categorized as non fragrant. Meanwhile, eight were found to be homozygous recessive and were categorized as fragrant. A total of 43 cultivars could not be distinguished due to problems in PCR optimization. Approximately, 34% of the genotyping results of the amplified samples were not parallel with the phenotypic results derived from the fragrance sensory evaluation test. This study provides a quick and easy way to identify fragrant rice variety which is valuable resources in crop development programs.