两种方法从林麝粪便提取DNA效率及分析

Analysis Two Methods for Extracting DNA from Musk Deer Feces

采集林麝DNA样本获得快捷、有效的DNA提取方法是其分子标记,良种遗传选育的分子基础。以林麝粪便为研究对象,用常规和改良的方法比较提取林麝粪便DNA的效率,两种方法分别设为对照组和试验组。采集林麝粪便后分别在空气暴露0、2、4、6d。用对照组和试验组两种方法提取粪便DNA,以林麝GAPDH为目的基因,用PCR方法扩增样本。琼脂糖凝胶电泳分离DNA目的条带,以空气暴露0d的粪便DNA条带光密度值为参考,2、4、6d粪便DNA条带光密度值与0d进行比较获得相对光密度值。结果发现,随着时间延长,林麝粪便逐渐失去光泽,表面粗糙;对照组和试验组两种方法均能有效提取林麝DNA。试验组2d和4d与0d相比较DNA水平显著上升,6d与0d没有显著差异;对照组仅在2d提取的DNA水平上升,而4d和6d显著下降到0d的水平。以对照组为参照,两组各时间点相比,试验组获得的DNA水平显著高于对照组。研究结果表明,试验组提取DNA的方法可显著提高林麝粪便DNA的获得率。

Molecular markers is a quick and efficient method for collecting forest musk deer DNA, and is also the fundament for selection of improved varieties. In this study, feces of forest musk deer was used as object, efficiency of DNA extraction from feces of forest musk deer was compared between normal and improved methods, and two methods were used as control and treated group, respective- ly. The feces were exposed for 0 d, 2 d, 4 d, 6 d after it was collected from forest musk deer. Two methods with control and treated group were carried out to extract feces DNA, and PCR was used to amplify samples by primer of GAPDH in forest musk deer. Target DNA fragment was separated by agarose gel electrophoresis. Optical density of DNA sample from feces exposed in air for 0 d for refer- ence, and relative optical density was obtained by comparing optical density from that of 2 d, 4 d and 6 d with that of 0 d. The results showed that feces were gradually tarnished and roughened with time- dependant. DNA was efficiently extracted from forest musk deer feces by both methods in control and treated groups. In treated groups, DNA level significantly increased by comparing 2 d and 4 d with 0 d, and there was no difference at 6 d, compared with 0 d; in control group, DNA level was only obvi- ously elevated at 2 d, compared with 0 d, and DNA level at 4 d and 6 d was declined to that of 0 d. With control groups used as reference, DNA level from treated group was significantly higher than that of control group at all time points. The results demonstrated that DNA could be efficiently ob- tained from forest musk deer feces from treated groups, which would provide the basis for molecular marker and breeding in forest musk deer.