Investigation Of NAL11 Regulating Flooding Tolerance In Rice Germination

Water direct seeding is an important sowing method of direct seeding rice.Because rice seeds are completely submerged in water,they suffer fromhypoxia or anoxia,which leads to poor germination or no germination.Therefore,breeding rice varieties with rapid germination and elongation of coleoptile under waterlogged conditions can effectively solve these limitations.Thus,the low oxygen germination is the key of rice direct seeding improvement andhas important significance to study on submergence tolerance of rice germination,improve and guarantee the yield of direct seeding rice,and promote its popularization.Studying NAL11,aheat shock protein identified earlier in our laboratory,the study was took the 02428 and its near-isogenic line 02428-nal11 as study materials to study the phenotype,spatio-temporal expression,physiological and biochemical characteristics of coleoptile under waterlogging stress,and the mechanism of NAL11 involved in ABA response to flooding in rice was also discussed.The main findings are as follows:1.Under waterlogged conditions,the coleoptile length and surface area of 02428-nal11were significantly longer and wider than 02428,but undifferentiated in the mean diameter.After phenotypic verification of the NAL11 knockout transgenic material of japonica rice variety ZH11,the results were consistent with those of 02428 and its near-isogenic line,In other words,the coleoptile length and surface area of ZH11^Ko-NAL11were significantly larger and wider than those of the control group,and no difference of mean diameter of ZH11Ko-NAL11.2.Comparing the expression of NAL11 germinates in air and waterlogged,we found thathypoxia can induce expression of NAL11.GUS staining analysis showed that GUS staining could be detected in coleoptiles germinated under air and waterlogged conditions,and the staining under the waterlogged was deeper than air.Besides,the GUS staining can express in the coleoptile axis.All these indicate that NAL11 could respond to adversity and express a lot in submerged germination.3.Underhypoxia stress,compared with 02428,the expression of amylase-related genes was up-regulated,the activity of amylase was increased,and glycolysis-related Adh1 and PDC1genes were up-regulated at 12 h and 48 h,that is,the activity of alcohol dehydrogenase and pyruvate dehydrogenase was increased.The results showed that nal11 could convert acetaldehyde that is toxic to cells underhypoxia stress into ethanol in time.It also provided favorable conditions for the rapid synthesis of ATP in rice coleoptiles underhypoxia stress and the maintenance of the p H value of cytoplasm.4.The results of stress-related enzyme gene quantitative analysis and enzyme activity under waterlogging stress showed that compared with 02428,the activity of CAT,SOD and POD in 02428-nal11 increased at 12 h and remained at ahigher level after 12 h.however,the content of MDA decreased with the increase of flooding time.These results indicated that nal11 could improve the scavenging ability of reactive oxygen species and decrease the degree of membrane lipid formation.5.Compared with 02428 and 02428-nal11,at 12 h,the expression of extended protein-related genes was down-regulated.But at 48 h,the expression of Os EXPA7 and Os EXPA10 were down-regulated,of Os EXPB7 were not significantly different,and of other related genes were up-regulated.6.Compared with 02428 and 02428-nal11,the expression of ABA synthesis genes in the coleoptile of 02428-nal1 was slightly down-regulated when waterlogging 12 h,and the catabolic genes were significantly increased.But the results were reversed at 48 h.The results of endogenoushormone ABA measurement showed that the ABA content in02428had no significant change in different flooding time,but increased obviously in02428-nal11 and showing a significant accumulation process.The results of quantitative analysis were consistent with the changes of endogenous ABA content.In addition,the expression of ABA signal transduction-related genes was down-regulated in 02428-nal11 at12 h after flooding,and then up-regulated in most of the genes at 48 h after flooding.It is suggested that NAL11 may be involved in ABA signaling and regulation of ABA response under flooding.