Cloning And Expression Vector Construction Of Phytochrome Genes PHYA And PHYB From Solanum Pinnatisectum

Light is an important factor in the plant life cycle. Receptors feeling light in plants include chlorophyll, thexanthophyl and phytochrome. Phytochrome (PHY) involve in many physiological processes such as plant seed germination, seedling growth, stem elongation, cotyledon extention and flowering control. There are five family members in PHYs, which are PHYA, PHYB, PHYC, PHYD and PHYE respectively. Some researches indicated that PHYA and PHYB also involved in the regulation of tuber formation and anthocyanins synthesis. But, their mechanism is not clear until now. In this research we cloned PHYA and PHYB genes from Solanum pinnatisectum by RT-PCR, analysed their expression in different organs and constructed their RNAi vectors, which provide working base for further study on the regulation mechanism of these two genes in tuber formation and anthocyanins synthesis. The results are as follows:The full-length cDNA of a gene PHYA was cloned from Wild Potato (Solanum pinnatisectum) by RT-PCR. The cDNA was 3 466bp long with a open reading frame (ORF) of 3 372 bp and encoded 1 123 amino acid residues with a predicted molecular weight of 125 kDa and an isoelectric point of 5.8. The protein sequence comparison showed that the predicted protein had higher identities to those from Solanum tuberosum, Lycopersicon esculentum and Nicotiana tabacum (96%, 94% and 91%). semi-quantitive RT-PCR indicated that the expression of PHYA was different depending on organs and higher in roots, stem and shoots. The influence of light on the expression of PHYA depended on the space position of organs and was promotive in the underground organ and restrained in the aboveground organs.The full-length cDNA of a gene PHYB was cloned from Wild Potato (Solanum pinnatisectum) by RT-PCR. The cDNA was 3470bp long with a open reading frame (ORF) of 3393 bp and encoded 1130 amino acid residues with a predicted molecular weight of 125 kDa and an isoelectric point of 5.6. The protein sequence comparison Showed that the predicted protein had higher identities to those from Solanum tuberosum, Lycopersicon esculentum and Nicotiana tabacum (98%,95%and92%). Semi-quantitive RT-PCR indicated that the expression of PHYB was similar among different organs, but little low in mature organs and independent of lighting and dark.In addition, in order to study the mechanism of PHYA in anthocyanins synthesis and PHYB in tuber formation two RNAi vectors pGSA1285PHYA and pGSA1285PHYB were constructed.