Screening Of Ginger Internal Reference Genes And Construction Of Ginger ZoARF2 Gene Vector

Ginger,as a kind of herb with the same origin of medicine and food,has high edible value and economic value.The study of some important functional genes in ginger is of great help to the growth and development of ginger.Real-time fluorescence quantitative PCR is an important method to analyze gene expression,and stable internal reference gene is a prerequisite for accurate quantification of gene expression level.Due to the imperfection of ginger database,stable internal reference genes have not been screened from ginger,so it is necessary to screen the internal reference genes of ginger.In this rearch,ginger was used as the research object,and qrt-pcr was used to study the expression of 11 candidate internal reference genes in different parts of ginger after postharment dehydration treatment.The expression stability of candidate internal reference genes and the number of introduced internal reference genes were determined after the analysis of geNorm,NormFinder and Bestkeeper,so as to better correct the expression analysis of ginger.After analysis,it was found that the two genes ACT2 and TUB2 could be used as internal reference genes for quantitative analysis of different parts of ginger,and three internal reference genes should be introduced for correction.After ginger water loss treatment,TUB2 is suitable as an internal reference gene for quantitative analysis to a certain extent,and it is recommended to use two combinations of internal reference genes as correction for quantitative analysis of gene expression.The results showed that TUB2 gene was the most suitable for quantitative analysis in all samples of ginger including different parts of ginger and water loss treatment of ginger.The edible part of ginger is subterranean fleshy rhizome,which is an important product organ and plays an important role in the formation of yield.In the reported species,ARF2 gene has been found to be a pleiotropic transcription factor that plays an important role in different growth and development stages of plants.From the transcriptomesequencing of early ginger,it was found that ARF2 gene was highly expressed in the process of rhizome enlargement,suggesting that ZoARF2 gene might play a regulatory role in the development of underground rhizome of ginger.In this study,the physicochemical properties of ZoARF2 protein in ginger were analyzed,and it was found that ZoARF2 protein was soluble but unstable.The ZoARF2 protein may be activated by amino acid sequence.According to the homologous relationship of amino acid sequence of ARF protein,it was found that it had a high homology with ARF2 amino acid sequence of Vitis vinifera L,Oryza sativa and Arabidopsis thaliana in the conserved region.Through qRT-PCR analysis,it was found that ZoARF2 gene in ginger was expressed in root,rhizome,stem and leaf,among which the expression in leaf was the highest,and the expression in rhizome was second only to that in leaf.There were some differences in expression between different tissues,suggesting that this gene may be involved in the different regulatory roles of ginger in different tissues.In addition,ZoARF2 expression vector has been successfully constructed in this experiment,which provides a foundation for further research on function of ZoARF2 gene.In summary,the results of this study provide important theoretical reference and technical support for the selection of internal reference genes and the analysis of gene expression in ginger,and lay a foundation for the follow-up studies on the function of ginger genes and the mechanism of root enlargement.