Cloning,Expression,Subcellular Localization And Genetic Transformation Of BZIP Transcription Factor Gene In Ramie

Ramie is a perennial fiber crops, belong to Urticaceae Boehmeria. Fiber yield and quality of ramie were seriously affected by abiotic stress. The production was severely reduced by summer and fall drought every year,; Therefore, It was very important that cloning, expression and characteristics analysis and genetic transformation the ramie stress related genes, further understanding the stress resistance molecular mechanism of ramie, and improvement the ramie varieties.In This study, the full length cDNA sequence of three bZIP transcription factor gene in XiangZhu3were cloned,and analysis organization expression specificity, stress induced expression and subcellular localization; Cloning and prediction the cis element of the portion of bZIP gene promoter sequence; The overexpression vector have been constructed, and obtained the BnbZIP1and BnbZIP3transgenosis Arabidopsis plants. The main results are as following:1. The full-length cDNA sequence and the ORF of BnbZIPl were2071bp and1407bp, which encoded468amino acids with predicted pI and molecule weight were4.95and36.81KD, respectively, and which contains bZIP transcription factor gene basic structure and typical three repeat leucine structure domain. Homology comparison analysis showed that the deduced BnbZIPl amino acid sequence was sharing78%、83℃、93%and72%homology with the bZIP gen with Soybean （NP₀01237098）、Tobacco （AAF06696）、 Western balsam poplar （XP₀02307972） and Butterfly orchid （ADK74339）, respectively. The results of subcellular localization analysis showed that the BnbZIPl located in nucleus. The results of real-time PCR suggested that the BnbZIPl gene expressed in root, stem, shoot tip and blade, female flowers and male flowers, with the highest expression level in male flowers and the lowest in root. The BnbZIPl gene was up-regulated by ABA, drought and high salt treatment. Cloning848bp the BnbZIPl gene ATG upstream promoter sequence, the online forecast analysis shows that its contain CAAT and TATA promoter core cis element, and also contains the drought induced, ethylene response cis function components, etc.; The pRI101-BnbZIPloverexpression vector was constructed, and import it into Agrobacterium, and obtained the transgenic Arabidopsis plants.2. The full-length cDNA sequence and the ORF of BnbZIPP2were1587bp and1416bp, which encoded471amino acids with predicted pi and molecule weight were7.73and52.19KD, respectively, and which contains bZIP transcription factor gene basic structure and typical three repeat leucine structure domain. Homology comparison analysis showed that the deduced BnbZIP2amino acid sequence was sharing73%、60%、74%and77%homology with the bZIP gen with Tobacco （AAY15214）、 Arabidopsis （CAC40649）、 Vitis vinifera （XP₀02275147） and Ricinus communis （XP₀02532393）, respectively. The results of subcellular localization analysis showed that the BnbZIP2located in nucleus. The results of real-time PCR suggested that the BnbZIP2gene expressed in root, stem, shoot tip and blade, female flowers and male flowers, with the highest expression level in female flowers and the lowest in stem. The BnbZIP2gene was up-regulated by ABA, drought and high salt treatment. Cloning796bp the BnbZIP2gene ATG upstream promoter sequence, the online forecast analysis shows that its contain CAAT and TATA promoter core cis element, and also contains the Gibberellic acid and salicylic acid response cis function components, etc.; The pRI101-BnbZIP2overexpression vector was constructed, and has imported it into Agrobacterium.3. The full-length cDNA sequence and the ORF of BnbZIP3were1756bp and999bp, which encoded332amino acids with predicted pI and molecule weight were9.16and37.22KD, respectively, and which contains bZIP transcription factor gene basic structure and typical three repeat leucine structure domain. Homology comparison analysis showed that the deduced BnbZIP3amino acid sequence was sharing91%、90%、82%and86%homology with the bZIP gen with Phaseolus vulgaris （AF402608）、Crabapple （ACU78078）、Rice （BAA72064） and Tobacco （AAF06696）, respectively. The results of subcellular localization analysis showed that the BnbZIP1located in nucleus. The results of real-time PCR suggested that the BnbZIP3gene expressed in root, stem, shoot tip and blade, female flowers and male flowers, and the expression difference is smaller. The BnbZIP3gene was up-regulated by ABA, drought and high salt treatment. Cloning671bp the BnbZIP3gene ATG upstream promoter sequence, the online forecast analysis shows that its contain CAAT and TATA promoter core cis element, and also contains the Methyl jasmonic acid and endosperm specific expression response cis function components, etc.; The pRI101-BnbZIP3overexpression vector was constructed, and import it into Agrobacterium, and obtained the transgenic Arabidopsis plants; Overexpression of BnbZIP3, which can improve the germination rate of transgenic arabidopsis under drought condition.