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针对SARS冠状病毒的分子生物学检测是控制SARS流行的关键环节。为评价全基因组扩增对SARS微量样本检测的影响 ,采用 6 mer随机引物反转录 ,用加接头的随机引物合成第二链 ,再以接头序列为引物扩增并掺入荧光标记 ,最后与带有 70 mer探针的基因芯片杂交。此非特异方法基本覆盖了样本中的全部DNA ,结果发现SARS冠状病毒全基因组的扩增效果对基因芯片杂交结果的均匀性有较大影响 ,PCR循环次数增多会导致扩增均匀性的降低。分析了不同的引物对全基因组扩增均匀性的影响 ,探讨了全基因组扩增策略的缺陷。
Molecular biology tests against SARS-CoV are key steps in controlling the SARS epidemic. To evaluate the effect of genome-wide amplification on the detection of SARS micro-samples, a reverse primer of 6 mer random primer was used to synthesize the second strand with a random primer with a linker. The second strand was amplified with the linker sequence as a primer and incorporated with a fluorescent marker. Finally, Gene chip hybridization with 70 mer probe. The nonspecific method basically covers all the DNA in the sample. The result shows that the genome-wide amplification effect of SARS coronavirus has a great influence on the homogeneity of the hybridization results of the gene chip, and the increase of the PCR cycles leads to the decrease of the uniformity of the amplification. The influence of different primers on genome-wide amplification was analyzed, and the defects of genome-wide amplification strategy were discussed.