Coleoptile Purple Line Regulated by A-P Gene System Is a Valuable Marker Trait for Seed Purity Identification in Hybrid Rice

In plants,a large number of anthocyanin biosynthetic genes encoding enzymes and regulatory genes encoding transcription factors are required for anthocyanin synthesis.Coleoptile purple lines are two purple lines on both sides of coleoptiles after seed germination.However,the molecular mechanism of coleoptile purple line is not clear in rice so far.In this study,two major dominant genes,coleoptile purple line 1(OsCPL1,also known as OsC1)and coleoptile purple line 2(OsCPL2),were isolated via map-based cloning,and both of them were required for anthocyanin biosynthesis of coleoptile purple line in rice.The knockout and complementation experiments confirmed that OsC1 was required for purple color in most organs,such as coleoptile line,sheath,auricle,stigma and apiculus,whereas OsCPL2 was just required for coleoptile purple line.OsC1 was predominantly expressed in coleoptiles,flag leaves,and green panicles,and highly expressed in young leaves,whereas OsCPL2 was predominantly expressed in coleoptiles,and extremely lowly expressed in the other tested organs.Loss-of-function of either OsC1 or OsCPL2 resulted in significant reduction of transcript levels of multiple anthocyanin biosynthesis genes in coleoptiles.Coleoptile purple line was further used as a marker trait in hybrid rice.Purity identification in hybrid rice seeds via coleoptile purple line just needed a little water,soil and a small plate and could be completed within 5 d.Molecular marker and field identification analyses indicated that coleoptile purple line was reliable for the hybrid seed purity identification.Our findings disclosed that coleoptile purple line in rice was regulated by two major dominant genes,OsC1 and OsCPL2,and can be used as a simple,rapid,accurate and economic marker trait for seed purity identification in hybrid rice.

A Simple SSR-based Method for Purity Identification of Cucurbita moschata Hybrids

Aiming at solving the existing issues in purity identification of Cucurbita moschata hybrids by SSR, such as complex operation and difficult application in production practice, in this study, a simple SSR-based method was established for purity identification of Cucurbita moschata hybrids. By using the established simple method, without grinding, freezing, centrifugation and drying, the genomic DNA extraction process is shorter than 3 min. Compared with the conventional method, PCR detection system and silver staining in polyacrylamide gel electrophoresis of the established method are more time-saving and cost-saving. The whole detection process is shorter than 4 h, and 480 samples can be detected with this method by one person in one day. In addition, the detection result exhibits a coincidence rate of 99% with field identification. The simple SSR-based method established in this study can provide basis for large-scale rapid purity identification of Cucurbita moschata hybrids.

Purity Identification of Xinkui 19 by SSR Marker Technique

In this study, by using Xinkui 19 and its parents as experimental materials, 100 pairs of SSR molecular markers were screened to obtain specific poly- morphic primers, aiming at providing an accurate and efficient method for identifying the purity of Helianthus annuus L. hybrid seeds. According to the experimen- tal results, by using polymorphic SSR primer 455, the amplified bands of female and male parents were 460 and 430 bp, respectively; by using polymorphic SSR primer 478, the amplified bands of female and male parents were 330 and 350 bp, respectively. The identification results of Xinkui 19 hybrids and its parents with SSR marker technique and field cultivation were basically consistent. SSR primers 455 and 478 could be used for rapid and effective identification of the purity of Xinkui 19 hybrid seeds.