Cloning And Genetic Transformation Of BZIP Transcription Factor In Bambusa Emeiensis

Bambusa emeiensis belongs to Bambusoideae Neosinocalamus, and it is dominant bamboo species in southwest China, with high cellulose content and good wood properties. But Bambusa emeiensis was influenced by freezing,drought and so on, which caused massive decline in its production. It was reported that bZIP transcription factors play an important role in response to abiotic stresses.In this research, six BebZIP genes were cloned based on the transcriptome database of shoots in Bambusa emeiensis and NCBI database. Their bioinformatics, transcriptional activity, tissue expression patterns, stress-induced expression analysis and transformation into Populus cathayana were performed. It provide a theoretical basis for improving the resistance of bamboo by transgenic engineering. The main results are as follows:Six BebZIP gene sequences were selected from the transcriptome and NCBI database. The full-length sequences of six BebZIP genes in Bambusa emeiensis were cloned. They were named BebZIP1, BebZIP2, BebZIP3, BebZIP4, BebZIP5 and BebZIP6, respectively. The full-length cDNA sequence of BebZIP1, BebZIP2,BebZIP3, BebZIP4, BebZIP5 and BebZIP6 were 1848 bp, 578 bp, 602 bp, 932 bp,894bp and 765 bp, and encoded 470, 167, 190, 292, 264 and 239 amino acids,respectively. Their accession numbers in GenBank were KU554692, KU554693,KU560570, KU560571, KU560572 and KU560573, respectively.It was found that the six bZIP amino acid sequences from Bambusa emeiensis contained highly conserved motif1. The six bZIP amino acid sequences included a bZIP superfamily. But the location of bZIP superfamily were not fixed in six bZIP amino acid sequences. Tertiary structure showed that six bZIP proteins were mainly composed of alpha helices. Tertiary structure of BebZIP2,BebZIP3, BebZIP5 and BebZIP6 protein had high similarity, and they were mainly composed of one or two alpha helixs. As for BebZIP1 and BebZIP4, their tertiary structure were relatively complex, which were composed of some alpha helixs and some random coils.The results of transcriptional activity analysis on six bZIP transcription factors from Bambusa emeiensis showed that the strong transcriptional activity was found in the transcription factor BebZIP1, BebZIP4 and BebZIP6, but BebZIP2 was weaker and BebZIP5 was weakest.The tissue expression patterns of six BebZIP genes were analyzed by the technique of real-time PCR. The results showed that six BebZIP gene sequences were expressed in all shoots, stems, unfolding leaves and rolled leaves of Bambusa emeiensis. Meanwhile, there were the expression differences of the same gene in the different tissues, but their overall performance were highest in the shoots.The seedlings of Bambusa emeiensis were treated with ABA(200 μmol ?L-1), NaCl(200 mmol ? L-1) and PEG 6000(20%) stress. The stress-induced expression in BebZIP2 and BebZIP6 genes was analyzed by Real-time PCR technology. The results showed that the expression level of BebZIP2 and BebZIP6 genes treated by salt, drought and ABA stress had different degree of sensitivity,respectively. We speculated that these two genes were both existed in metabolic pathways of salt, drought and ABA stress, thus they affected the response of Bambusa emeiensis to abiotic stress.The plant overexpression vectors named pCAMBIA1303-BebZIP6 was successfully constructed, and it was successfully transformed into Populus cathayana. The transgenic Populus cathayana plant was obtained, which laid a foundation for functional research of BebZIP6 gene in the future.