Development and utilization of the functional co-dominant KASP marker for thermo-sensitive genic male sterility in rice Oryza sativa L.

Abstract

Thermo-sensitive genic male sterility (TGMS) is an important genetic means of two-line hybrid rice breeding. Pollen fertility in TGMS lines is regulated by a single point mutation in TGMS genes. Based on single nucleotide polymorphisms (SNPs) and insertion-deletion mutations, Kompetitive Allele-Specific PCR (KASP) markers were developed and utilized in rice molecular breeding via high-throughput detection, which saves time and money. In this study, we converted the SNPs on TGMS genes (including p/tms12-1and tms9-1) to functional co-dominant KASP markers and bred the resultant two-line hybrid rice. We differentiated the TGMS lines carrying p/tms12-1 or tms9-1 from other TGMS lines using the KASP assay. Pei’ai64S and Hua201S (containing p/tms12-1) had a homozygous GG genotype generating a blue signal. HengnongS-1 (containing tms9-1) had a homozygous CC genotype generating a red signal. The KASP assay for tms9-1 was identified as a recessive Mendelian trait. Seed purity was tested using the KASP marker for the two-line hybrid varieties of Liangyoupeijiu and Hualiangyou1206, which was consistent with findings by the dCAPS marker. Moreover, new TGMS lines were generated by pyramiding tms12-1 and tms9-1 gene under the same genetic background. Therefore, the KASP marker to detect the TGMS genes developed in this study can be widely used in two-line hybrid rice breeding. It provides a visually convenient toolkit for breeders to select individual target plants using high-throughput screening in two-line rice breeding.