Genetic variation at the waxy locus associated with starch pasting properties in international rice germplasm

Authors: Chen, Ming Hsuan; Bergman, Christine; Fjellstrom, Robert

Submitted to: Rice Technical Working Group Meeting Proceedings
Publication Type: Proceedings
Publication Acceptance Date: 12/1/2003
Publication Date: 6/1/2004
Citation: Chen, M., Bergman, C.J., Fjellstrom, R.G. 2004. Genetic variation at the waxy locus associated with starch pasting properties in international rice germplasm. In: Proceedings of the 30th Rice Technical Working Group Meeting. 2004 CDROM.

Interpretive Summary:

Technical Abstract: Rice starch pasting properties are used to characterize the processing and cooking qualities of rice (Oryza sativa L.). The Rapid Visco Analyser (RVA) has become the standard method for determination of rice pasting properties. The Waxy gene on rice chromosome 6 encodes the granule-bound starch synthase enzyme, which controls much of the variation in grain amylose content, and reportedly has major effects on starch pasting properties. Amylose content, however, does not explain all the variation in rice pasting characteristics. Some varieties with similar apparent amylose content have very different pasting viscosities. Rexmont, a javanica type rice with a strong pasting curve, has a characteristic single nucleotide substitution resulting in an amino acid change in exon 10 of the Waxy gene (ex10 SNP). We are studying 164 diverse rice accessions with apparent amylose contents ranging from 0 to 27%, and determining the sequence variation in the Waxy gene to identify associations between pasting properties and genotypes and its linkage to the Waxy CT-repeats microsatellite marker. Genomic DNA of rice varieties were extracted from leaf tissues. The CT repeat was determined using the Waxy microsatellite marker. The sequence variation in exon 10 was detected by the dideoxyfingerprinting method, a hybrid between single-strand conformation polymorphism and the dideoxy sequencing method, and was scored against a Nipponbare control. The rice pasting properties were determined by RVA. Two allele-types of the ex10 SNP, namely the RXMT-type allele associated with a strong RVA curve (S-RVA allele), and the NPBR-, or Jodon-type, allele associated with a weak RVA curve (W-RVA allele), were identified from 164 rice accessions including non-glutinous and glutinous varieties. A mean comparison of the individual characteristics of the RVA curve demonstrated that the S-RVA allele has significantly (alpha = 0.05) higher hot paste viscosity, cool paste viscosity, and setback than those of the W-RVA allele regardless of the amylose content. A genetic marker, via allele-specific oligonucleotide PCR (ex10-ASO), to this ex10 SNP has been developed and is able to discriminate a strong RVA curve of the RXMT-type from a weak RVA curve of the NPBR-, or Jodon-type. In addition, this S-RVA allele is linked to Waxy CT 11 repeats, suggesting that both ex10-ASO and the Waxy microsatellite marker are suitable to be used in breeding programs for varietal development of pasting properties. However, amylose content did contribute to the pasting properties of the RVA curve of the rice accessions with the W-RVA allele, excluding the glutinous varieties. The amylose content is negatively correlated with the RVA of peak viscosity, hot paste viscosity, and breakdown (r = -0.85***, -0.75*** and -0.79***, respectively), and is positively correlated with the setback (r = 0.76***). In conclusion, the high association of ex10-SNP and Waxy CT repeats to rice starch pasting properties as observed in the evaluation of the diverse international germplasm suggests these ex10-ASO and Waxy microsatellite markers can be used as markers in varietal development programs using international accessions. Since RVA is environment dependent, and requires 3 grams of flour from mature rice kernels, the availability of markers will accelerate and increase accuracy in progeny selection in the breeding program.