Study Of The Alternative Splicing Genes And Resistance To High Temerature In Populus Tomentosa

High temperature weather appeared more and more frequently,which had a great impact on the growth of plants.In this study,Populus tomentosa was used as the material and treated at four temperatures of 25℃,30℃,35℃ and 40℃.Through transcripts sequencing,the gene alternative splicing(AS)events and the relation to heat resistance were revealed.The main results are as follows:(1)The physiological indexes of Populus tomentosa leaves were measured at 25 ℃,30 ℃,35 ℃ and 40 ℃.The results showed that at 25 ℃,30 ℃ and 35 ℃,the content of proline,malondialdehyde,conductivity,soluble sugar and chlorophyll did no significant change,but when the temperature increased to 40 ℃,the content of proline,malondialdehyde,conductivity and soluble sugar increased significantly,while the content of chlorophyll decreased significantly.It can be seen that 25 ℃,30 ℃ and 35 ℃ have no significant effect on Populus tomentosa,while 40 ℃ has significant effect on Populus tomentosa.(2)A total of 10777,11605,10680,and 14577 AS events were separately identified under the treatment of 25℃,30℃,35℃,and 40℃,which related to 4105,4276,4079,and 4915 genes,respectively.Of them,the intron retention pattern was the most common,accounting for42.31%～51% of all AS events,and exon skipping pattern was the least,accounting for9.14%～10.23% only.chromosomes 9 had the most condensed AS events of 107.03 events/Mb while chromosome 19 had the least of 51.25 events/Mb.Association analysis showed that AS was positively correlated with the intron length,exon number,exon length,and gene expression level,and was negatively correlated with GC content.(3)A total of 2001 AS genes under the treatment 40℃ were identified in comparison to that of25℃.GO and KEGG enrichment analysis showed them mainly involved in the RNA degradation pathway and valine,leucine and isoleucine degradation pathway.The results showed that AS,as an important way of post-transcriptional regulation,not only participated in the process of plant growth and development,but also had a positive impact on stress resistance of plants.(4)The expression analysis showed that Ptt EXPA11 had a normal splicing transcript of Ptt EXPA11-cds at 25℃,30℃ and 35℃,while had AS occurrence produced two transcript isoforms of Ptt EXPA11-L and Ptt EXPA11-S at 40℃.Both of the isoforms were confirmed not to be degraded by the NMD pathway,and possibly encoded a single domain protein.All the three transcripts were cloned and assembled to the vetor of p EZR(K)-LC.Transformation to tobacco plants was conducted,some regenerated seedlings were obtained for the next function evaluation to heat stress.