Functional Analysis Of The HOOKLESS1 Gene In Arabidopsis And Preparation Antibody Of Its Encoded Protein

To quickly break through the soil and touch the light to carry out photosynthesis,dicotyledon form an apical hook structure to protect meristems from damage when they germinated in the dark.The formation of the apical hook is affected by hormonal,gibberellin and ethylene and other environmental conditions such as light.Some report indicated that ethylene response gene HOOKLESS1 is also involved in the regulation of the formation of apical hooks.The Arabidopsis HLS1 is related to the regulation of auxin distribution and expression of glycoresponsive genes and plant immunity.In this study,the HLS1 gene was used as the research object,and its mutants hls1-1 and Salk₁36528c were used as materials to study the function of HLS1 gene.We have obtained the following experimental results:?1?T2 transgenic plants of different environments were stained by GUS histochemistry,and the results showed that HLS1 was mainly expressed in new tissues and organs in plants.Subcellular localization of HLS1 by Arabidopsis WT mesophyll cell protoplast transformation experiments revealed that HLS1 is localized in the nucleus and late endosomes,we deducing the HLS1 is related to some biological pathways that about endosomes.Phylogenetic tree and protein sequence alignment analysis found that the biological function of HLS1 is highly conserved.?2?The HLS1 gene mutants hls1-1 and Salk₁36528c were used as the genetic background.A binary expression vector was constructed and through Agrobacterium transformation method overexpressed HLS1 to establish complementation lines.Studies on the complementation lines found that overexpression HLS1 could complement the hls1-1 and Salk₁36528c apical hooks deficiency and the early flower phenotype to wild type.The results of the carbon starvation induced aging showed that the premature aging phenotype of the mutant also recovered,indicating that HLS1 may be involved in the regulation of plant leaf senescence.?3?We constructed the prokaryotic expression vector pET28a-HLS1,followed by transformation into BL21?DE3?E.coli.After IPTG induction,HLS1-His was purified via Nickel affinity chromatography.HLS1-His antiserum was obtained through immunning rabbits.After affinity purification of antibody,we identified the accumulation of HLS1 by HLS1 antibody in transgenic plants.The specific antibody against HLS1 protein was prepared successfully,which laid a foundation for studing the function of HLS1 in plant development.