Construction The Plant Experessing Vector Of Transcription Factors StPti5 From Solanum Tuberosum And Analysis Its Stress-induced Experssion Characteristics

The life cycle of plant can't be in suitable environment conditions completely,abiotic and biotic stress will company the whole life.The expression of transcription factors in adversity conditions can improve the resistance of plants to the stress environment,and play a very important role in the long time coevolution of plants and the environment.AP2/EREBP(APETALA2/ethylene-res ponsive element binding proteins)is a plant specific transcription factor superfamily.The ERF transcriptional factor is a subgroup of AP2/EREBP transcription factors,whose main function is to regulate the response of plants to hormones(ethylene and ABA),pathogens and stress(low temperature,drought and high salt).Early transcriptome sequencing results show that expression level of a ERF gene PGSC0003DMG400010309 named StPti5 of Jiaxiang 1 was up-regulated significantly after inoculation of Phytophthora infestans 24 hours.We cloned the gene and constructed the plant expression vector and analysis the sequence of the gene for studying the function of the gene.Real-time fluorescence-based quantitative RT-PCR were carried out of StPti5 under stress conditions such as P.infestans inoculation,high temperature,low temperature,drought and high salt stresses.The main results as follow:1.Gene cloning and sequence analysis: primers of Jiaxiang 1's StPti5 were designed according to the sequences of PGSC0003DMG400010309 gene and the primer 5 'end add the sequence of BamHI and SalI enzyme sites.The StPti5 gene was obtained by RT-PCR from the RNA of J Jiaxiang 1.The largest open reading frame(ORF)of StPti5 is 468 bp and encodes 156 amino acids.The bioinformatics analysis of amino acid sequence of the gene and the sequence of other ERF transcription factor amino acid sequences show that the gene contains the AP2 domain,which plays an important role in adversity.2.Plant expression vector construction and transformation into Agrobacterium: the StPti5 was cloned into expression vector pCAMBIA1301 m.StPti5 gene had been cloned into expression vector by sequencing and enzyme digestion verification,which named p1301m-StPti5.Next we transform the p1301m-StPti5 into the Agrobacterium GV3101 byheat shock.This work provides the basis for function study of this gene.We obtained 4positive plants of Arabidopsis thaliana by hygromycin plate screening and PCR detection.3.Real-time fluorescence-based quantitative RT-PCR analysis of StPti5 gene: single element such as the P.infestans,high temperature,low temperature,drought stress and high salt were tested,we found the StPti5 gene was up-regulated after inoculation P.infestans48h-72 h.StPti5 gene was down-regulated at 4?.The high temperature of 35?,the high salt of 150mmol/L NaCl and the simulated drought treatment of 20%PEG,the express level of StPti5 first rose and then declined.