果汁DNA提取方法比较及柑橘属植物分子生物学检测技术的研究

Comparison on the Extraction Method of DNA and Study on the Biotechnological Detection of Orange Component in Juice

以橙汁为研究对象,选取市售100%橙汁及橙汁饮料各3种,采用CTAB法、SDS-CTAB法以及改良CTAB法提取其DNA,对所提取的DNA的浓度、纯度及扩增效率进行比较;同时选取橙UDP-葡糖基转移酶蛋白基因设计柑橘属植物特异性扩增引物,通过定性PCR检测其特异性和灵敏度,并应用于果汁中柑橘属植物的检测。实验结果证明,由改良CTAB法获得的DNA模板质量较高,适用于果汁中DNA的提取。UGT基因引物可对柑橘属植物产生特异性扩增,最低检测限为10pg/μL。将该引用物用于市售橙汁的检测,其检测结果为阳性,从而证明UGT基因特异性引物可用于果汁中的柑橘属植物成分的检测及鉴伪研究。

Three methods, including CTAB Method, SDS-CTAB method and modified CTAB method, were used to extract DNA from both pure orange juice and juice beverage, then the quality and quantity of DNA were analyzed by spectrophotometer and PCR amplification; the orange-specific primers were designed according to the sequence of UDP- glucoxyltransferase protein gene. The results showed that modified CTAB method was more suitable for the juice DNA extraction than the other two methods; the orange component could be detected by the orange-specific primers with a limit of detection of 10 pg/μL. The following detection of fruit juice samples showed that this method was suitable to identified the orange component in fruit juice and other relative food and can provide a technique base for research of fruit juice adulteration detection.