Promoter Analysis Of TaMOC1 Gene And Obtaining Of The CRISPR-Cas9 Mutants In Triticum Aestivum L.

Wheat?Triticum aestivum L.?is the main food crop,and is widely cultivated in the world.Tiller is closely related to yield,and is one of the most important agricultural traits of wheat,so it is important to understand the molecular mechanism of tillering in wheat.The MOC1gene is the key gene that controls the occurrence and growth of tiller buds in rice.In previous study,we have cloned the TaMOC1 gene and made a preliminary study of its expression pattern and function.Gene expression regulation contains the transcriptional level regulation,post-transcriptional regulation,translational regulation,etc.The transcriptional level regulation is important,and the promoter plays a vital role in transcriptional level regulation.Therefore,it is important to study the sequence structure and action elements of promoter for understanding the gene expression pattern.In this study,we cloned the promoter sequence of TaMOC1 and preliminarily analyzed its sequence structure and action elements.In addition,to confirm the function of TaMOC1 in wheat,the TaMOC1 CRISPR/Cas vector was transfected into wheat?cultivated variety KN199?.The results are as follows:?1?The semi-quantitative PCR analysis showed that the expression patterns of TaMOC1-7A?TaMOC1-7B and TaMOC1-7D were expressed in roots,shoot apexes,leaves and spikelets.TaMOC1-7B was higher expressered in the shoot apexes.?2?The cis-acting element analyses of the promoters of TaMOC1-7A?TaMOC1-7B?TaMOC1-7D show that TaMOC1 promoters might be induced by hormones,and be involved in multiple stress signaling pathways.The meristem expression related cis regulatory elements might play an important role in regulating the expression of TaMOC1.?3?GUS staining results showed the CAT-box regulatory element migtht determins the expression pattern of TaMOC1 gene.?4?Five pBUE411-TaMOC1-Cas9 T0 generation transgenic wheat plants were obtained,and their seed rate were lower.It suggested that the mutation of TaMOC1 might affect the development of spliklets.The sequencing analysis of T1 generation transgenic plants showed that partial nucleotide and amino acid sequences had been changed in the target point sequence.The expression levels of TaMOC1 in pBUE411-TaMOC1-Cas9 transgenic wheat T0 generation plants were also decreased than that in WT control.Next,we will detect the GUS staining signals in T2 generation P_TaMOC1-7D::GUS wheat plant to understand the expression pattern of TaMOC1.At the same time,we will observe the total tiller phenotypes of pBUE411-TaMOC1-Cas9 transgenic wheat plants in the field.