Mutations in wheat starch synthase II genes and PCR-based selection of a SGP-1 null line

Abstract

Wheat (Triticum aestivum L.) starch synthase II, which is also known as starch granule protein 1 (SGP-1), plays a major role in endosperm starch synthesis. The three SGP-1 proteins, SGP-A1, B1 and D1, are produced by three homoeologous SSII genes, wSSII-A, B, and D. Lines carrying null alleles for each SGP-1 protein have previously been identified. In this report, the mutations occurring in each wSSII gene were characterized, and PCR-based DNA markers capable of detecting the mutations were developed. In the null wSSII-A allele, a 289 bp deletion accompanied by 8 bp of filler DNA was present near the initiation codon. A 175 bp insertion occurred in exon 8 of the null wSSII-B allele. The insertion represented a recently discovered miniature inverted-repeat transposable element (MITE) named Hikkoshi that was first found in a wheat waxy gene. A 63 bp deletion was found at the region surrounding the junction of the fifth exon and intron of the null wSSII-D allele. Based on this information, we designed primer sets to enable us to conduct allele-specific amplifications for each locus. The applicability of these primer sets for breeding programs was demonstrated by reconstructing a line lacking all three SGP-1 proteins using marker-assisted selection. These markers will also be useful in breeding programs aimed at obtaining partial mutants missing one or two SGP-1 proteins.

Sweet Wheat

“…The materials and methods used in this study are as follows: Plant materials included the wild-type cultivar Chinese Spring (CS), the Wx common wheat cultivar Mochi-Otome and a HA mutant line developed in our previous work (Shimbata et al, 2005). Double mutants were selected by PCR-based screening of the F 2 and F 3 generations of a cross between the HA and Wx mutant lines with six primer sets (Nakamura et al, 2002;Shimbata et al, 2005; Table 2).…”

“…Double mutants were selected by PCR-based screening of the F 2 and F 3 generations of a cross between the HA and Wx mutant lines with six primer sets (Nakamura et al, 2002;Shimbata et al, 2005; Table 2). Seeds were harvested at 45-50 days post anthesis (DPA) for seed morphology observations and at 25 DPA for sugar content measurements.…”

Starch Modification: A Model for Wheat MAS Breeding

“…After SSIIa-A1 , − B1 and -D1 null lines were identifi ed, these lines were crossed and a fully null SSIIa mutant was selected. The amylose content in this line was increased by about one-third as compared to wild-type Shimbata et al 2005 ). Although no commercially available HA variety has been released yet, quality studies have shown dramatic changes in fl our properties for this type of wheat Shimbata et al 2005 ).…”

“…The amylose content in this line was increased by about one-third as compared to wild-type Shimbata et al 2005 ). Although no commercially available HA variety has been released yet, quality studies have shown dramatic changes in fl our properties for this type of wheat Shimbata et al 2005 ). HA seed also contains a higher proportion of resistant starch (Yamamori et al 2006 ), which is thought to have a beneficial infl uence on health.…”

“…Again, the use of MAS appeared to be the most effi cient way to obtain the desired genotypes. We fi rst characterized the mutations in the three SSIIa genes at the DNA sequence level, then used this information to develop PCR-based markers that were co-dominant and could be multiplexed (Shimbata et al 2005 ). After reconstituting the HA line by MAS, we used MAS to develop near-isogenic lines of the eight possible homozygous genotypes for the SSIIa gene, using a leading variety from the southern area of Japan as the recurrent parent.…”