| Picea wilsonii Mast.,also known as North China Spruce,is an evergreen coniferous tree of Pinaceae.It is a unique tree species in China.It has a beautiful tree shape and plays an important role in northern landscaping.The ecological value.In recent years,with the appearance of various environmental problems,abiotic stress factors such as drought,salinity,and extreme temperature have seriously threatened the habitat of the indigo.Therefore,it is particularly important to improve the resistance of the indica by various genetic strategies.The use of molecular biology methods to clone and study the stress-resistance related genes in the indica cristata can effectively protect high-quality functional genes in woody plants,enrich the theoretical basis of plant stress resistance research,and provide genetic resources for stress-resistant genetic engineering of forest trees.With theoretical and technical support,we will breed new varieties of anti-retroviral genes to protect the species of barley.In this study,the anti-stress gene screening and functional validation of alfalfa were tested,and some genes in the transcription system library of alfalfa were screened using fluorescence quantitative analysis technology.After the target gene was obtained,bioinformatics analysis and expression patterns under stress were analyzed.Plant genes expression analysis in different organs,subcellular localization and transformation of the target genes in Arabidopsis thaliana have been experimentally studied and validated.The main research results are as follows:1.Eleven Candidate Genes Responsive to Salt and Drought Stress were screened out from the transcript profile of Picea wilsonii and verified by RT-qPCR experiments.The results showed that the expression levels of PwGS,PwCFE,PwH2A and PwHb under salt and drought stress conditions are most significant.2.To predict the function of four genes with significant difference in expression,PwGS protein is involved in the plant oligosaccharide synthesis pathway,PwCFE is involved in the synthesis of plant cell fibrin,PwH2A is a subunit of chromatin histone,PwHb is the plant Hemoglobin,involved in plant oxidative stress.3.Bioinformatics analysis of the target genes,in which PwGS,PwH2A and PwHb proteins are structurally stable,are hydrophilic proteins,no signal peptides,do not contain transmembrane domains,are located outside the membrane,and are both in the secondary structure.Including a-helix,extended chain structure and random coiling,the ratio of the three is equal;PwCFE protein is unstable,and the disordered features are obvious.The secondary structure mainly contains a-helices and random coils.Subcellular localization predictions indicate that PwGS and PwHb are localized in the cytoplasm and PwH2A and PwCFE are located in the nucleus and mitochondria,respectively.4.RT-qPCR results showed that PwGS was responsive to ABA,salt,and drought,and was highly expressed in the stems and leaves of the barley;PwH2A responded significantly to heat stress and was mainly expressed in the leaf tissue of the indigo;PwHb to ABA The response to salt stress was significant,and it was abundantly expressed in the leaves of indigo.PwCFE showed specific response to salt and drought stress,and the highest expression in leaves.5.The results of subcellular localization experiments showed that PwHb is mainly located in the cytoplasm and PwH2A is mainly located in the nucleus,which is consistent with the software prediction.6.PwH2A and PwHb were transfected into A.thaliana by Agrobacterium-mediated transformation and TO transgenic positive plants were obtained. |
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