利用EST计划获得的1β680个基因片段,制备了国内外首张茶树cDNA芯片,芯片上每个基因设置一个重复,共含有6β912个点,其中6β720个靶基因,160个阳性对照,32个阴性对照。4×4矩阵点制,共两个区域,每个区域的芯片密度为1β037点/cm2,每张芯片可进行两次杂交。用3个茶多酚含量有差异的品种与芯片进行了杂交,获得了不同品种的基因表达谱及表达差异的基因;选取其中2个与茶叶香气密切相关的基因,采用荧光定量PCR的方法进行了验证,基因表达变化趋势与芯片检测结果一致,验证了cDNA芯片杂交结果的可靠性。该芯片可以进一步应用于许多茶学研究领域,进行基因表达差异的高通量检测。
Totally, 1β680 genes obtained in our EST project were selected from the cDNA library of clone Longjing 43 to develop the first cDNA microarray of tea plant. Each gene in the microarray was duplicated. The cDNA microarray contains 6β912 dots, including 6β720 EST, 160 positive controls and 32 negative controls. One microarray was spotted two regions with 4×4 matrixes and it could be hybridized twice. The density of the microarray was 1037 dots per cm2. Three tea clones with different contents of tea polyphenol were selected to conduct two sets of hybridization experiments with the microarray and the differently expressed ESTs were found. Then, two genes that were closely related to tea aroma were selected for real-time PCR analysis to validate the microarray data and validated the reliability of the cDNA microarray. The newly developed cDNA microarray could be applied in various tea research fields to make the high-throughput detection in the gene expression profiling.
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