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The Puroindoline b-2 Variants are Expressed at Low Levels Relative to the Puroindoline D1 Genes in Wheat Seeds
- M. J. Giroux, K.-H. Kim, A. C. Hogg, J. M. Martin, B. Beecher
- Crop science 2013 v.53 no.3 pp. 833-841
- RNA, Triticum aestivum, chromosome elimination, cultivars, gene expression, genes, genetic variation, genotype, hardness, loci, seeds, sequence analysis, starch granules, transcription (genetics), wheat
- Grain hardness in wheat is largely controlled by the Hardness locus. This locus contains the Puroindoline a (Pina) and b (Pinb) genes, thought to exist as single-copy genes only on chromosome 5D. However, four additional copies of Pinb have been reported. These new genes termed Pinb-2v1 to Pinb-2v4 map to the group 7 chromosomes of wheat (Triticum aestivum L.). Since PINA and PINB cooperatively bind to starch granules and PINB limits grain softness, we set out to determine Pinb-2 gene variant expression levels relative to Pinb. The tested genotypes included the soft wheat cultivar Alpowa, Chinese Spring, and two Chinese Spring group 7 nullisomic-tetrasomic lines. Deletion of chromosome 7A (and Pinb-2v4) or 7D (and Pinb-2v1) did not impact either the level of total or starch-bound puroindolines, indicating that the expression level of the Pinb-2 genes is not sufficient to modify grain hardness. Due to cross-hybridization, northern blots were not useful in accurately quantifying Pinb-2 transcript levels. Therefore, RNA sequencing was used to accurately measure transcript levels of Pina, Pinb, and each of the four Pinb-2 variants in developing wheat seeds. Relative to Pinb, Pinb-2v1 expression levels averaged 1%, whereas Pinb-2v2 levels ranged up to 7.6%. Pinb-2v3 transcripts were detectable only in Alpowa at 1% of Pinb-D1a, whereas Pinb-2v4 transcripts were undetectable in all genotypes. Given the relative low expression levels of each Pinb-2, it is unlikely that they substantially contribute to grain hardness variation.