Development and validation of CAPS-marker associated with the Rf2 gene in sorghum (Sorghum bicolor (L.) Moench)

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Abstract
Background. The development of heterotic hybrids based on cytoplasmic male sterility (CMS) is the leading strategy in breeding sorghum (Sorghum bicolor (L.) Moench). The trait of pollen fertility restoration in forms with CMS A1 (milo), predominantly used in sorghum breeding, is determined by at least two dominant complementary genes Rf1 and Rf2, and also gene Rf5. The development of accessible molecular markers of sorghum Rf genes is highly relevant for hybrid breeding, since they can significantly accelerate the process of creating female sterile forms (A lines), sterility maintainers (B lines) and pollen fertility restorers (R lines).Material and methods. The studied material included 36 sorghum accessions from the VIR collection, which differed by the ability to restore pollen fertility in forms with A1-type CMS. The nucleotide polymorphism of 935 bp fragments of the PPR genes Sobic.002G057050, Sobic.002G054100, and Sobic.002G054200 located at the chromosome 2 was studied.Results. The fragments obtained with the use of a pair of 2459403fw and 2459403 primers were 935 bp long and included parts of three genes: Sobic.002G057050, Sobic.002G054100, Sobic.002G054200. For identifying the sequence variant Sobic.002G057050-1090 associated with the Rf2 gene, Tru9 I restrictase was chosen, which allows obtaining a 572 bp fragment unique for all the studied R lines. Such a marker was found in 10 sorghum lines from West China and Kyrgyzstan, which are widely used in breeding as fertility restorers. The fragment was found neither in three lines with sterile cytoplasm and their fertile analogues, nor in 7 accessions of kafir sorghum, which lacked functional alleles of Rf genes.Conclusions. It has been demonstrated that the marker can be used for selection and checking purity of R and B/A lines. It is also applicable for verifying hybridity of F1 seeds and analyzing hybrid populations from crosses of R lines 924-4, 928-1, 929-3, 931-1, 933-1/6, 1237-3, 1243-2, 1251, 1150-1, F10BC2 with A lines Nizkorosloe 81s, А-83 and А-10598. It may be suggested that the ability to restore pollen fertility in R lines, which lack the marker CAPS- 572, is determined by another Rf gene. The studied 935 bp fragment of Sobic.002G057050 harbours 22 SNP, therefore the development of CAPS-markers for their identification and differentiation can be promising.

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