Black rice has high accumulation of anthocyanins on pericarp tissues. OsB1 and OsC1 genes are important for the controls of anthocyanin biosynthesis in rice. OsDFR gene is structural gene that encodes the vital enzyme in anthocyanin biosynthesis pathway. Our research granted in the year of 2019 showed that DNA markers specific to OsB1, OsDFR and OSC1 genes could be used to distinguish between white and black rice. In this study, we used these DNA markers to study the F2 population of 300 plants derived from the cross between white pericarp rice (Pathumthani 1) as a receptor and black pericarp rice (Kham Noi) as a donor. The inheritance of DNA markers specific to OsB1, OsDFR and OSC1 genes was analyzed by Chi-square test with 1, 2 and 3 genes. The results showed that DNA markers specific to OsB1 and OsDFR followed the Mendelian manner but DNA marker specific to OsC1 did not. Analysis of anthocyanin content by pH differential method in seeds of F2 progenies showed that plant no. 299 had highest anthocyanin content of 35.04 mg/100 gDW, consistent with 39.52 mg/100 gDW of the cyanidin-3-O-glucoside (C3G) content determined by HPLC method. Antioxidant activity was analyzed by DPPH assay in seeds of F2 progenies and the result showed that plant no. 199 had highest antioxidant activity of 9.7 ?moles/gDW which was similar to that of Kham Noi parent. Analysis of inheritance of F3 pericarp colors by Chi-square test showed the ratio of 3 colored: 1 colorless, corresponding to genotype of OsB1 marker of 3 A_: 1 aa. The relationship between genotype of DNA markers specific to OsB1, OsDFR and OsC1 and phenotype of F2 population was analyzed by ANOVA and regression method. The results suggested that only OsB1 marker was related to anthocyanin content, antioxidant activity and pericarp color which had R2 of 31.9 %, 40.6 % and 69.5 %, respectively.
The inheritance of DNA markers specific to OsB1, OsDFR and OSC1 genes was individually analyzed by Chi-square test in 321 plants of BC1F1 population. The results showed that only DNA marker specific to OsB1 followed the Mendelian law which had the ratio of 1:1 (Aa:aa). The 15 plants of BC1F1 population which were heterozygous for DNA markers of OsB1, OsDFR and OSC1 genes were selected. They had different pericarp colors including 8 black, 4 dark brown, 2 dark red and 1 red which were further backcrossed with Pathumthani 1 for production of BC2F1 seeds and selfing for production of BC1F2 seeds. Antioxidant activity was analyzed by DPPH assay in seeds of BC1F1 and the result showed that plant no. 297 had highest antioxidant activity of 12.8 ?moles/gDW which was higher than 8.8 ?moles/gDW of Kham Noi parent.
In this study, the CAPS marker of OsB1 had relationship with phenotype of anthocyanin content, antioxidant activity and pericarp color. Therefore, it could be used for selection of rice with colored pericarp, high anthocyanin and antioxidant activity in the early stage of development which will be beneficial to plant breeding for facilitating selection and shortening the time in breeding Thai rice for high nutritional value. In addition, the seeds of BC2F1 and BC1F2 which were selected by the DNA markers and had black and red pericarp colors, high anthocyanin and antioxidant activity, will be further used for improvement of rice varieties with increased nutritional value.