GENE EXPRESSION ANALYSIS OF ISOFLAVONE SYNTHESIS PATHWAY ON F4 GENERATION FOR STRAIN EVALUATION IN LOCAL BLACK SOYBEAN (Glycine max. (L.) Merr)

• Main Author: Marwa Mumtazah, Aulia
• Format: Final Project
• Language: Indonesia
• Subjects: Ilmu hayati ; Biologi
• Online Access: https://digilib.itb.ac.id/gdl/view/40042
• Institution: Institut Teknologi Bandung

Black soybean (Glycine max. (L.) Merr) is an important agricultural commodity in Indonesia for the production of food, drugs, and chemicals.. Black soybeans contains isoflavone 100 times greater compared to other legumes. Isoflavones are secondary metabolites that play a role in plant defense mechanisms and have many benefits for human health. The synthesis of isoflavone is controlled by several genes and environmental factors. Information on the function of these genes controlling isoflavone synthesis is needed in the use of molecular markers for the development of black soybean varieties which have characteristics of high and stable isoflavones. In this study, the expression of genes involved in isoflavone biosynthesis pathway (CHS8, CHR, CHI1A, F6H3, and IFS2) was profiled on local varieties of UP106 and UP122 black soybeans and F4 generation of UP106xUP122 and UP122xUP106 to evaluate potential strain lines for cultivar selection. Both F1 strains have relatively stable isoflavone synthesis with significant difference (UP106 high levels of isoflavone and UP122 low levels of isoflavones) and F4 generation which is the result of the two F1 reciprocal crosses, namely UP106xUP122 and UP122xUP106. The research methods includes planting, sampling, isolation of RNA, quantity and quality analysis of RNA, synthesis of cDNA, reverse transcriptase PCR (RT-PCR) CHS8, CHR, CHI1A, F6H3, and IFS2 genes. The expression of those genes was semi-quantified based on band thickness from agarose gel electrophoresis using ImageJ. Relative expression were quantified against actin (ACT2/7) as reference gene, The results showed that in the relative expression level of CHS8 and CHR were highest in the UP106 genotype, expression of CHI1A was not significantly different between all genotypes based on one-way ANOVA test (p-value>0.05). IFS2 expression was highest in F4 genotype (UP106xUP122. Expression analysis of F6H3 gene was not conducted due to the absence of bands from RT-PCR results. The conclusion of this study is that F4 generation (UP106xUP122) shows highest IFS2 expression than both F1 strains. IFS2 is one of the key genes in isoflavone biosynthesis. In addition, the profile of gene expression in both generations of F4 was influenced by crossing UP106xUP122 and UP122xUP106 on CHS8, CHR and IFS2 genes based on t-test (p-value <0.05) and had no effect on CHI1A and F6H3 genes.