Cloning And Function Identification Of Stress Resistance-related NAC Transcription Factors From Miscanthus Lutarioriparius (Poaceae)

Miscanthus is a kind of perennial rhizomatous grasses, which has high biomass productivity, extensive adaptability and low input cost. Now, the research about Miscanthus focuse on breeding new variety which can grow on marginal land, such as saline-alkali soil and desert. As the plant-specified transcription factor, NAC involved in multiple biology processes of plant, including development, lateral root formation, hormone regulation, senescence, and abiotic and hormone stress responsive etc. This study intent to find NAC genes which have the potential ability to improve Miscanthus’stress tolerance in Miscanthus lutarioriparius. By sequence analysis, expression profile analysis, functional identification by transgenic Arabiodopsis etc, we analyzed some stress-related NAC transcription factors in Miscanthus lutarioriparius.1. Full-length cDNA sequences were obtained for11MINAC genes, and we conducted sequence alignment, phylogenetical analysis and domain prediction. The sequence alignment result indicates that the N-terminal region of NAC proteins was highly conserved, and the11deduced proteins were further divided into five NAC subdomains (A-E). Conversely, the C-terminal transcription activation region was extremely diversified, In this study, domain3,8,1,2, and10specified the NAM subdomain A to E, respectively. Some specific domains were identified in specific subfamilies, for instance, domain7,9, and12for subfamily OsNAC3, domain14for subfamily ATAF, and domain6for subfamily ONAC003. These11MlNAC transcription factors were phylogenetically classified into six subfamilies. As the NAC members exhibited an interspersed distribution in each subfamily, it could be inferred that the expansion of NAC genes occurred before the divergence of Miscanthus, Arabidopsis, and rice.2. We carried out quantitative real time RT-PCR (RT-qPCR) to monitor13MINAC genes’ expression profiles in different tissues and under different abiotic stress and hormone treatment. The results show that some MINACs predominantly expressed in some tissues and some MINACs’ expression was detected in all above six tissues. All surveyed13MINACs were up-regulated under salt, drought, MeJA and SA treatment; under cold stress,7MINACs were up-regulated and3MINACs were down-regulated; under ABA treatment, MINAC4and MlNAC10were down-regulated, the others were up-regulated; the expression of all genes surveyed except MlNACll showed significant increase in wounding treatment.3. We also conducted subcellular localization and transcriptional activation analysis of MlNAC2, MlNAC5and MlNAC13. They all localized in nucleus and had transcriptional activation in full-length and C-terminal region. Overexpressed MlNAC2by CaMV35S in Arabiodopsis, the transgenic plant’ drought tolerance was improved. And the specific function of MlNAC2needs further verification.