Improvement Of Drought Tolerance Of Switchgrass By Overexpressing One Aquaporin Gene PvPIP2;9

Switchgrass(Panicum virgatum)is a perennial warm-season grass in the grass family(Poaceae).Switchgrass is featured with its tall statue,rapid growth,robust root system,and high resistance to most stresses and adaptive to adverse environment.It has been used as the non-grain-based bioenergy plant as well as the soil and water conservation plant.Identification the resistance genes and applying such knowledge onto molecular breeding of switchgrass and the other related Poaceae species is a promising way to further improve its stress resistance for the purpose of restoration,improvement and utilization of marginal land.Plant aquaporins play an important role in maintaining the water status in plants.PIP is one subfamily of the aquaporins.The PIP proteins affect the ability of plants to take up water and some other small molecules.A number of the PIP subfamily genes can positively regulate plant abiotic stress,especially the adaptation to drought stress.In this study,we cloned and analyzed a switchgrass PIP gene,PvPIP2;9,gene and found that over-expressing the PvPIP2;9 gene significantly improved the drought resistance of switchgrass.The results of this study are as follows:(1)Phylogenetic tree analysis showed that PvPIP2;9 is close to OsPIP2;4,OsPIP2;7,and ZmPIP2;7 in rice and maize.The result of multiple sequence alignment show that PvPIP2;9 as well as its orthologous proteins had the typical 6 transmembrane domains and two conservative NPA motifs in the aquaporin family.(2)The expression pattern of PvPIP2;9 was analyzed.The results showed that PvPIP2;9 was regulated by the circadian rhythm and its expression fluctuated along with the photopeirod.The expression level increased after illumination and reached its maximum after 4 hours,and then decreased.The expression of PvPIP2;9 was also up-regulated by cold and drought treatment,but down-regulated with salt and ABA treatment.(3)Using Agrobacterium-mediated genetic transformation,we generated PvPIP2;9 overexpression transgenic switchgrass lines.The transgenic and wild-type switchgrass were compared for their drought tolerance.We found that the drought tolerance of the transgenic switchgrass was significantly stronger than that of wild-type switchgrass.The related physiological indexes of plant drought tolerance were measured.During the water withholding period,the EL of transgenic switchgrass became significantly lower than that of the wild-type,while the leaf RWC of transgenic switchgrass was significantly higher that of the wild-type,these show that transgenic switchgrass could maintain better cell membrane integrity and higher moisture content to deal with drought stress.We also found that the transgenic switchgrass maintained higher Fv/Fm and chlorophyll contents than those of the wild-type,indicating transgenic switchgrass could maintain better photosynthesis to maintain normal development of plants under the drought stress conditions.Furthermore,we also found that the WUE of transgenic switchgrass were significantly higher than that of wild-type.In conclusion:the expression of PvPIP2;9 gene was affected by the circadian rhythm,and was affected by different abiotic stress factors.Overexpression of PvPIP2;9 gene in switchgrass can improved leaf WUE and photosynthesis under drought treatment and alleviated the damage of cell membrane,thus enhancing the drought tolerance of switchgrass.