Effects Of Enhanced UV-B Radiation On AtFH16 Of Arabidopsis

Plants are fixed organisms that need to regulate their growth to adapt to changing environments.Light is essential for photosynthesis.It can also be used as a source of information about the environment,seasons or time of day,enabling plants to adjust their morphology,development or metabolism.UV-B radiation(280-315 nm)is located at the far end of the short-wave radiation in the solar spectrum,and the energy of 290-315 nm reaches the surface of the earth.The stratospheric ozone depletion is accompanied by the increase of UV-B radiation,which has attracted wide attention from the scientific community at the end of the 20 th century.Formin proteins are highly conserved multi-domain proteins that interact with both microfilaments and microtubules.When wheat seedlings were exposed to enhanced UV-B radiation,it was found that chromosomes aggregated into 3-6 bundles and moved toward the cell poles,accompanied by polynucleation.This special phenomenon of mitosis is called“bundle splitting”.Previous studies have shown that AtFH16 may regulate cell division by regulating the behavior of microtubules,microfilament aggregation and bunching.The results obtained in this paper are as follows:(1)AtFH16 gene was cloned using cDNA as template,and the expression vector35S-AtFH16-GFP was successfully constructed.(2)Using DNA as template,the promoter fragment pAtFH16 of AtFH16 gene was cloned,and the CaMV35 S constitutive promoter of pCAMBIA1300-GFP was successfully constructed,and the expression vector pFH16-AtFH16-GFP was successfully constructed.(3)The transcriptional level of AtFH16 in fh16 mutant was identified.AtFH16 gene was not expressed in fh16 mutant,and fh16 was a homozygous mutant.(4)The 35S-AtFH16-GFP expression vector was transformed into wild-type byimmersion method,and the pFH16-AtFH16-GFP expression vector was transformed into fh16 mutants and wild-type.Transgenic positive plants were screened by hygromycin.Finally,homozygous overexpression plants of 35S-AtFH16-GFP,homozygous restorer plants of pFH16-AtFH16-GFP and homozygous overexpression plants of pFH16-AtFH16-GFP were obtained.(5)Phenotypic observation of wild type,fh16 mutants and transgenic plants showed that the plant height,root length and leaf area of fh16 mutants were lower than those of wild-type;35S?AtFH16/WT and pFH16?AtFH16/WT were higher than those of wild-type in plant height,root length and leaf area.The plant height,root length and leaf area of pFH16?AtFH16/fh16 were the same as those of wild type,which could restore the phenotype of fh16 mutants.After enhanced UV-B radiation,the expression of AtFH16 decreased.Micronuclei were observed in root tip cells of fh16 mutant after UV-B treatment.(6)In addition,wheat is an important food crop,and there is no reports about wheat UVR8 at present.In this paper,the common wheat cultivar ‘Jinmai 47' was studied,and the UVR8 gene sequence was obtained by RT-PCR.The characteristics of the wheat UV-B photoreceptor UVR8 gene and its encoded protein were systematically analyzed by bioinformatics.Results showed a 1329 bp sequence that encodes 442 amino acids corresponding to wheat UVR8.The gene was named TaUVR8 and submitted to GenBank with the accession number of MK170136.Conserved domain analysis predicted that the protein encoded by the TaUVR8 gene contains seven RCC1 domains,similar to AtUVR8 in Arabidopsis thaliana.Phylogenetic tree analysis showed that TaUVR8 and BdUVR8 proteins were clustered in the same branch,and their genetic distances were relatively close,indicating that they had a close evolutionary relationship.3-D structural analysis indicated that the protein is constituted by a seven-bladed ?-propeller,which has a high structural homology with AtUVR8 in Arabidopsis thaliana.This study provides a theoretical basis for studying the molecular mechanisms of UV-B photomorphogenesis and the function of the wheat TaUVR8 gene.