Cloning Of A YEATS Domain-Containing Gene OsYAF9 And Creating Material In Rice

Flowering time is one of the most important agronomic traits for regional and seasonal adaption in rice.The Heading stage is affected by both endogenous and exogenous factors.Although the molecular basis of flowering time control is well dissected in the long day(LD)plant Arabidopsis,it is still largely unknown in the short day(SD)plant rice.During the last years,many evidences have shown that the YEATS domain is evolutionarily conserved.It's found in more than 100 different eukaryotic proteins and participate in transcriptional and chromatin modifications.In yeast,Yaf9,Taf14 and Sas5 constitute the YEATS domain protein family.Among them,Yaf9 is a subunit of the histone H4 acyltransferase complex Nu A4,and is also a subunit of the chromatin remodeling complex SWR1,which mediates the polymerization and separation of the Nu A4 and SWR1 complexes,thereby affecting its function.The YEATS domain is required for the Yaf9 driving function.When the yeast is deficient in Yaf9,it is sensitive to DNA damage and cold damage.The yeast Yaf9 protein has two homologs in Arabidopsis,YAF9 a and YAF9b(TAF14),and its YEATS functional domains are sequence-conserved in yeast,rice,alfalfa,human and fruit flies.The study found that YAF9 a and YAF9 b are redundant in function,and the double mutants exhibit multiple phenotypic defects which include early flowering.Among them,YAF9 a controls flowering time by regulating the level of FLC histone H4 acetylation.However,the function of the the gene contained in the rice YEATS domain has not been reported so far.In order to study the function of the YEATS domain-containing gene in rice,we used reverse genetics method to gain a homologous gene OsYAF9 which is the YEATS domain-containing gene.Evolutionary analysis and conservation analysis of the functional domain,the results showed that the sequence and functional domains of this gene were evolutionarily conserved in different species.At the same time,with the CRISPR-Cas9 and DNA recombination technology,we constructed the overexpression and knockout vectors of OsYAF9,and transformed rice variety SN9816 with rice genetic transformation technology,and obtained the T1 transgenic plants;we transient transformation of rice protoplasts with the 35 S promoter-initiated CDs and fusion of GFP expression vector,and conducted the subcellular localization analysis.The results showed that the protein encoded by OsYAF9 gene was localized in the nucleus.We used the real-time PCR method to detect the expression characteristics of OsYAF9 in different tissues and organs of rice.The results showed that OsYAF9 gene was constitutively expressed in roots,stems,leaves,sheaths,growth point, young ears of 1-3cm,5-7cm,and 7-13 cm,and ears of 4 days after fertilization,7 days after fertilization.In summary,we analyzed the evolution and domain's conservation of OsYAF9 gene in rice by bioinformatics,and created gene knockout and overexpression materials which is necessary for studying gene function,and laid the foundation for further study of the function of the OsYAF9 gene.