Mining And Expression Analysis Of Transcription Factors Of Citrus Plasma Membrane Proton Pump Gene CsPH8

The low expression of CsPH8,a plasma membrane type proton pump gene,is an important reason for low acid character of HL(Citrus sinensis cv.Honganliu).In this experiment,the homologous genes of AN1,AN11,PH3 and PH4 were cloned in citrus by mining orange genome database and using AL(C.sinensis cv.Anliu)and HL as the research materials.T hese genes were analyzed with bioinformatics method.The tissue-specific analysis and the expression dynamics during the development of fruits of these genes were studied,as well as the relationship between these genes and the change of citric acid content and gene expression of CsPH8.The roles of these genes in regulation of CsPH8 expression and citric acid accumulation were preliminarily identified.The results are as follows:1.The change of citric acid content and expression analysis of CsPH8 during citrus fruit development.This study found that the content of citric acid in the fruit of AL and ZXY(C.grandis cv.Zaoxiangyou)showed a trend of rise first then fall in the course of fruit development,while the content of citric acid in the fruit of HL and AFP(C.grandis cv.Acid free Pumelo)was significantly lower than that of AL and ZXY and had no obvious change.The CsPH8 expression in the fruit juice of AL and ZXY juice sac was increased first and then decreased,while the expression of CsPH8 in HL and AFP juice sac remained at a lower level and was significantly lower than that of AL and ZXY.2.Mining and Bioinformatics analysis of homologous genes of AN1,AN11,PH3 and PH4A AN1 homologous gene(named CsAN1),a AN11 homologous gene(named CsAN11),a PH3 homologous gene,which contains three transcripts(named CsPH3-1,CsPH3-2,CsPH3-3)and a PH4 homologous gene(named CsPH4)were cloned from the citrus genome,respectively.The proteins encoded by these genes are bHLH transcription factors,WD40 transcription factors,WRKY transcription factors and MYB transcription factors,respectively.3.Tissue-specific analysis of AN1,AN11,PH3 and PH4 homologous genesThe expression of AN1,AN11,PH3 and PH4 homologous genes in different tissues of AL was determined.It is found that the expression level of CsAN1 in juice sac was the highest.The expression level of CsAN11 in leaves was the highest.The expression level of CsPH3-1 in flowers was the highest.The expression level of CsPH3-2 in stems was the highest.The expression level of CsPH3-3 in leaves and stems was the highest.The expression level of CsPH4 in stem was the highest.4.Expression analysis of AN1,AN11,PH3 and PH4 homologous genes during fruit developmentIt was found that the expression of CsAN1 gene in AL and ZXY was significantly higher than that in HL and AFP,and the change trend of gene expression was the same as that of citric acid in AL and ZXY,and the same trend as CsPH8 gene expression.Thetrend of CsAN11 gene expression was not the same as that of citric acid in AL and ZXY.The expression levels of CsPH3-1 gene in AL and ZXY were significantly higher than that in HL and AFP,and the trend of gene expression was the same as that of citric acid in AL and ZXY.On the other hand,the trend of CsPH3-2 and CsPH3-3gene expression was different from that of citric acid in AL and ZXY.At the same time,the expression levels of CsPH3-1 in the all periods of citrus fruit of AL and HL were significantly higher than those of CsPH3-2 and CsPH3-3.The expression level of CsPH4 gene in the late development of AL and ZXY was significantly higher than that of HL and AFP,and the trend of gene expression was the same as that of citric acid in HL and AFP.Based on the above results,we speculated that the low expression of CsPH4 and CsPH3-1,especially CsAN1 in HL,may be an important reason for the low expression of CsPH8 in HL.They may be involved in the regulation of low acid content in HL by regulating CsPH8.