Screening and functional analysis of potential <i>S</i> genes in <i>Chrysanthemum morifolium</i>

Abstract

<i>S</i> genes are the key genes that cause plant self-incompatibility, to find out the key <i>S</i> genes and understand molecular mechanism of self-incompatibility in chrysanthemum, the stigmas and anthers at different developmental stages of 'Q10-22-2'—a self-incompatible chrysanthemum cultivar, were used for RNA sequencing. After bioinformatics analysis, 13 candidate pistil <i>S</i> genes and five candidate pollen <i>S</i> genes were excavated. A potential pistil <i>S</i> gene was cloned and named as <i>CmSRK1</i>. Meanwhile, a potential pollen <i>S</i> gene was cloned and named as <i>CmPCP1</i>. qRT-PCR revealed that <i>CmSRK1</i> was specifically expressed in mature stigmas, and <i>CmPCP1</i> was specifically expressed in anthers 3 d before maturation. Subcellular localization showed that both CmSRK1 and CmPCP1 were located in the nucleus and the cell membrane. Transcriptional activation activity analysis indicated that both of the two proteins had no transcriptional activation activity. Yeast two hybrid assay showed that there was no interaction between CmSRK1 and CmPCP1. <i>CmSRK1</i> was constructed on the expression vector containing stigma-specific promoter, and <i>CmPCP1</i> was constructed on the expression vector containing pollen-specific promoter, they were then transformed into <i>Arabidopsis thaliana</i>. Artificial hybridization was performed with transgenic lines containing <i>CmSRK1</i> as the female parents, and transgenic lines containing <i>CmPCP1</i> as the male parents. The hybridization results showed that seed sets of two transgenic lines were 19.62% and 11.64%, respectively, while cross-pollinated seed sets of Col-0 was 84.43%. Therefore, it was speculated that <i>CmSRK1</i> and <i>CmPCP1</i> might be pistil and pollen <i>S</i> genes of chrysanthemum, respectively, and SI of chrysanthemum belonged to SSI.