Allelic variation of the β-, γ- and δ-kafirin genes in diverse Sorghum genotypes

Abstract

The β-, γ- and δ-kafirin genes were sequenced from 35 Sorghum genotypes to investigate the allelic diversity of seed storage proteins. A range of grain sorghums, including inbred parents from internationally diverse breeding programs and landraces, and three wild Sorghum relatives were selected to encompass an extensive array of improved and unimproved germplasm in the Eusorghum. A single locus exists for each of the expressed kafirin-encoding genes, unlike the multigenic α-kafirins. Significant diversity was found for each locus, with the cysteine-rich β-kafirin having four alleles, including the first natural null mutant reported for this prolamin subfamily. This allele contains a frame shift insertion at +206 resulting in a premature stop codon. SDS-PAGE revealed that lines with this allele do not produce β-kafirin. An analysis of flour viscosity reveals that these β-kafirin null lines have a difference in grain quality, with significantly lower viscosity observed over the entire Rapid ViscoAnalyser time course. There was less diversity at the protein level within the cysteine-rich γ-kafirin, with only two alleles in the cultivated sorghums. There were only two alleles for the δ-kafirin locus among the S. bicolor germplasm, with one allele encoding ten extra amino acids, of which five were methionine residues, with an additional methionine resulting from a nucleotide substitution. This longer allele encodes a protein with 19.1% methionine. The Asian species, S. propinquum, had distinct alleles for all three kafirin genes. We found no evidence for selection on the three kafirin genes during sorghum domestication even though the δ-kafirin locus displayed comparatively low genetic variation. This study has identified genetic diversity in all single copy seed storage protein genes, including a null mutant for β-kafirin in Sorghum.

Allelic variation at a single gene increases food value in a drought-tolerant staple cereal

“…Plant leaf samples were collected fresh or stored dried and extracted with a CTAB buffer method as described by Laidlaw et al 28 . PCR for genotyping was carried out with Bioline MyTaq HS (www.bioline.com.au) per the manufacturer's protocol.…”

Grain Proteomics of Sorghum: Improving Digestibility, Nutritional Quality and Starch Conversion to Ethanol

“…1D separation of the prolamin fraction revealed altered expression of β-kafirin in a 19kD protein band, present in 296B and absent in QL12 (Fig. 3.4, highlighted in white), as predicted based on previous molecular characterisation (167). On 2D gels differential expression of β-kafirin was identified in 296B…”

“…Twenty eight inbred sorghum lines, with previously determined allelic variation in the β-, γ-, and δ-kafirin storage proteins were included in the study (167). The primary focus of the analysis was on comparisons between the β-kafirin null line QL12 and in 296B, an important food line of Indian origin.…”

Breeding strategies to improve sorghum quality

“…These markers were mainly used for genetic diversity, cultivar identification, gene mapping and discovery, and gene pyramiding. For instance, The β-, γ-and δ-kafirin genes were sequenced from 35 sorghum genotypes to investigate the allelic diversity of seed storage proteins (Laidlaw et al, 2010). Marker-assisted selection (MAS) is becoming an increasingly useful tool to breeders, and it is our goal to add them to the toolbox by providing a genetic analysis of sorghum protein digestibility.…”

“…The γ-kafirin contains 9-12% of the total kafirin fraction and 19-21% of opaque endosperm, whereas the σ-kafirin contains 1% total seed storage protein. Laidlaw et al (2010) reported characterization of alleles for presence of genetic diversity among the seed storage proteins in sorghum cultivars. The sorghum cultivars were sequenced and characterised for grain protein quality where the β-kafirin showed large variation in grain quality, and cysteine rich γ-kafirin and σ-kafirin comprised less variation.…”