Cloning and Expression Analysis of PeNAC047 Gene from Phyllostachys edulis
ZHAO Zhong-Yu1, HOU Dan1, HU Qiu-Tao1, WEI Han-Tian1, ZHENG Ying1, LIN Xin-Chun1,2,*
1 State Key Laboratory of Subtropical Silviculture, Zhejiang A & F University, Lin'an 311300, China; 2 Zhejiang Provincial Collaborative Innovation Center for Bamboo Resources and High-efficiency Utilization, Lin'an 311300, China
Abstract Moso bamboo (Phyllostachys edulis) is the most important economic bamboo species in China. Drought, salinity and other stress not only affected the yield of bamboo, but also limited its distribution. NAC (NAM, ATAF1/2 and CUC2) is one of the largest transcription factor families in plants, which plays an important role in the plant growth, development and responding to stress. In this study, in order to explore the molecular mechanism in the abiotic stress, this study cloned the PeNAC047-1 (GenBankNo. MN013397) and PeNAC047-2 (GenBankNo. MN013398) which were the homologous gene of ANAC047. The phylogenetic tree of PeNAC047-1 and PeNAC047-2 proteins were constructed by MEGA7.0 software. Multiple sequence alignment of PeNAC047-1 and PeNAC047-2 proteins were analyzed by DNAMan. The gene structure was analyzed with NCBI Conserved Domains Database (CDD) and the acting elements in upstream regulatory sequence were detected using PlantPan 2.0 online software, respectively. Transcription factor characteristics were verified by subcellular localization and transcriptional activation experiments. The expression pattern of PeNAC047-1 and PeNAC047-2 in different tissue and abiotic stress including abscisic acid (ABA), methyl jasmonate (MeJA), gibberellin acid 3 (GA3), drought and NaCl were analyzed by qRT-PCR. The result showed that the coding regions of the PeNAC047-1 and PeNAC047-2 were 879 bp and 1 095 bp, encoding 292 and 364 amino acids, respectively. The regulatory sequence upstream of PeNAC047-1 and PeNAC047-2 were 2 000 bp, which were including many kinds of responsive elements such as GA responsive element, MeJA responsive element, ABA responsive element etc. It's indicated that PeNAC047-1 and PeNAC047-2 might be regulated by hormones and abiotic stress. Based on subcellular localization and transcriptional activation experiment assay PeNAC047-1 and PeNAC047-2 was shown to be the nuclear protein and transcriptional activator. Tissue specific expression indicated that PeNAC047-1 and PeNAC047-2 transcripts gene were expressed in all tissues of bamboo, among which the highest level of PeNAC047-1 and PeNAC047-2 was found in root. Besides, PeNAC047-1 and PeNAC047-2 was induced by high salt. Under drought stress, the expression of PeNAC047-1 and PeNAC047-2 were inhibited, but the degree of inhibition of PeNAC047-1 was weakened, and PeNAC047-2 was inhibited continuously. The expression of PeNAC047-1 and PeNAC047-2 was induced by high temperature and the expression pattern was consistent basically. Similarly, the expression of PeNAC047-1 and PeNAC047-2 was induced by MeJA and the expression pattern was consistent basically. After ABA treatment, the expression pattern of PeNAC047-1 and PeNAC047-2 were different, PeNAC047-1 was induced by ABA, and PeNAC047-2 was inhibited. In addition, under the GA3 treatment, the expression patterns of PeNAC047-1 and PeNAC047-2 were also different. The response of PeNAC047-1 to GA3 was earlier (3~6 h), and the response of PeNAC047-2 to GA3 was later (48~72 h). In conclusion, PeNAC047-1 and PeNAC047-2 had the highest expression levels in root and respond to abiotic stress. And then the expression patterns of high salt, drought, high temperature and MeJA treatment were similar. Specially, the response of PeNAC047-1 and PeNAC047-2 with ABA and GA3 were inconsistent, so, it was speculated that it might be respond to abiotic stress by different pathways. This study explored the response pattern of PeNAC047-1 and PeNAC047-2 with abiotic stress preliminarily, which provides some basis data for the molecular mechanism of Moso bamboo in the abiotic stress.
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