Using Cherry Chalcone Synthase CpCHS1 Gene To Create Tobacco Drought-Resistant Germplas

’Manaohong’ cherry(Cerasus pseudoceresus ’Manaohong’)belongs to Chinese cherry,which is a cherry variety with excellent quality,storage and transportation resistance selected independently in Guizhou Province.The planting area has exceeded 20,000 hectares.As a key development tree in Guizhou’s 14 th Five-Year Plan,’Manaohong’ cherry will play an important role in Guizhou’s rural revitalization.The transcriptome analysis of ’Manaohong’ cherry root system revealed that the expression of Chalcone Synthase(CHS),a key enzyme of flavonoid synthesis and metabolism pathway in cherry root system,was significantly up-regulated under the adverse conditions.The gene was significantly up-regulated,suggesting that this gene may be closely related to the high resistance of ’Manaohong’ cherry.In this study,we first cloned one CHS gene of ’Manaohong’ cherry,named CpCHS1,and performed bioinformatics analysis of its sequence.The CpCHS1 gene was genetically transformed into tobacco using Agrobacterium-mediated method,and the transgenic strain was identified for drought resistance,which laid the foundation for the utilization of the gene.The main findings are as follows.1.Snapgene software was used to design specific primers for cloning the coding region of the gene(CpCHS1-F: 5’-ATGGTGACCGTGGAGGAAGTTC-3’CpCHS1-R:5’-TCAAGCGTTCAAGCCCACG-3’).The full length of the coding region was amplified by PCR using ’Manaohong’ cherry c DNA as the template.’Manaohong’ cherry was 1227 bp in length,encoding 408 amino acids.The relative molecular mass was 44.6 k D,and the theoretical isoelectric point(p I)was 6.53;there was no transmembrane structural domain,and it was a stable hydrophilic protein;the protein signal peptide predicted that the gene was a non-secretory protein;the secondary structure included α-helix and irregular curl,etc.;multiple sequence comparison showed that CpCHS1 was most closely related to peach(Prunus persica),and the similarity was 100%.2.qPCR showed that the expression of CpCHS1 was highest in roots,followed by fruits and leaves.3,The plant overexpression vector was constructed for the gene,and nine stably heterologously expressed transgenic tobacco strains were obtained by genetic transformation of tobacco using the leaf disc method.4,Drought stress treatment was applied to seeds,seedlings and adult seedlings of trans-CpCHS1 gene tobacco respectively,and it was found that the heterologous overexpression of CpCHS1 promoted the germination rate of seeds;both root length and fresh weight of tobacco seedlings were increased,and the drought resistance of tobacco was significantly enhanced,these results indicated that trans-CpCHS1 gene tobacco provides the basis for drought-resistant germplasm of tobacco.