Cloning and expression analysis of S-RNase homologous gene in Citrus reticulata Blanco cv. Wuzishatangju.

ABSTRACT

S-RNase-based self-incompatibility is the most widespread form of genetically controlled mate selection in plants and that S-RNase controls pollination specificity in the pistils. 'Wuzishatangju' (Citrus reticulata Blanco), a nature bud mutant from a self-compatible (SC) cultivar 'Shatangju', displays gametophytic self-incompatibility (GSI). In this study, full-length sequences of cDNA and DNA of the S-RNase homologous gene were obtained from 'Wuzishatangju' and 'Shatangju'. There was no difference in ORF sequences of the S-RNase cDNA between 'Wuzishatangju' and 'Shatangju'. However, 13, 9 and 6 consecutive bases were missing in 'Wuzishatangju' cDNA 5' UTR, 3' UTR and genomic DNA, respectively. Tissue-specific expression of the S-RNase gene was detected using semi-quantitative RT-PCR and quantitative real-time PCR. The expression level of the S-RNase gene in styles of 'Wuzishatangju' was approximately 10- and 5-fold higher than that in leaves and pollen, respectively. When 'Wuzishatangju' was self-pollinated, the expression of S-RNase in pistils peaked at 3 days, which was approximately 10-fold higher than that at 4h and 7 days, while in cross-pollination of 'Wuzishatangju' x 'Shatangju' the expression was very weak at 3 days. Results from a Southern blot showed that two copies of the S-RNase gene existed in genomic DNA of both 'Wuzishatangju' and 'Shatangju'.