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Citrus greening disease (Huanglongbing, HLB) is caused by the citrus greening bacteria (Ca. Liberibacter) citrus devastating disease germs as well as citrus and Huanglong Huanglong germs infection process of interaction mechanism remains unclear, citrus Huanglongbing anti- disease breeding is still no major breakthrough. Research citrus host of the the Huanglong pathogen response mechanism to help elucidate the pathogenesis of disease to develop effective management measures. The purpose of this study is based on the pre-build the citrus healthy plants infected Huanglongbing diseased subtractive (SSH) library was screened differentially expressed CsMYB (MYB transcription factor), they CsCAD (cinnamyl alcohol dehydrogenase) gene disease has been EST sequences by cloning and analysis of the function of these genes, expression using quantitative real-time PCR technology to track and study them the role of the Huanglong bacteria infection explore the citrus pathogen infection mechanism and citrus infected with Huanglong germs defense mechanism. Anti-citrus Huanglongbing molecular biology studies provide a basis for the citrus disease prevention and citrus breeding for resistance to provide a theoretical basis and technical support, so that the further development of citrus disease prevention and treatment. In this study, the method according to CsMYB EST sequences of primers were designed in CsCAD RNA was extracted from the leaves of sweet orange (Citrus sinensis) rapid amplification of cDNA ends the technology (RACE) were cloned CsMYB, CsCAD gene full-length cDNA, while taking advantage of the bioinformatics prediction analysis of the features and functions of the two genes; using real-time quantitative PCR analysis of two gene expression characteristics of in Huanglong bacteria stress. The major findings and conclusions of this study are as follows: 1. Received CsMYB (GenBank Accession No. HQ841074) full-length cDNA and bioinformatics analysis. Full-length cDNA of CsMYB gene 1306bp, bioinformatics analysis, including a 909bp complete open reading frame of the gene as well as a typical 26bp poly-A, encoding 302 amino acids, molecular weight 32.97kD, isoelectric point of 8.5 Meanwhile, there are conserved features MYB gene region, i.e. at the N-terminus, there are two typical MYB DNA binding domain: R2 and R3; obtained CsCAD (GenBank Accession No. HQ841075) cDNA full-length, full-length for 1124bp. Bioinformatic analysis shows the gene, including a 897bp complete open reading frame encoding 298 amino acids molecular weight of 32.45kD, and an isoelectric point of 6.4, while there are two conserved the region: ADH_N ADH_zinc_N. Using quantitative PCR analysis CsMYB and CsCAD expression in different periods of Huanglong pathogen infection. The quantitative analysis shows CsMYB gene in different periods of infection, CsMYB gene expression presents the trend of \Compared to the control, about a month or so after inoculation, CsMYB gene expression level is low, a weak upward trend; month later, along with the proliferation of bacteria, the amount of gene expression is increased to a higher level, as the control of the 2-3 times citrus Huanglongbing disease appeared three months later; Later, as the disease worsens, amount of CsMYB gene expression began to fall even lower levels than the control, and maintained at a low level. The gene expression CsCAD also basic show the trend of \In about 20 days after inoculation, CsCAD expression basically did not change, but the expression levels rose rapidly after a month, up to 10 times more than three months later, the expression levels began to decline, and, ultimately, to maintain at a lower level , but still higher than that of the control. 4. Cloned CsMYB, cDNA sequence of CsCAD, analysis of the features and functions of their sequence, according to the quantitative analysis of the results obtained at the same time, presumably CsMYB is a transcription factor gene, CsCAD is cinnamyl alcohol dehydrogenase gene, they may be involved in the citrus Huanglong pathogen defense response process.
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