Cano R J, Poinar H N
Biological Sciences Department, California Polytechnic State University, San Luis Obispo 93407.
Biotechniques. 1993 Sep;15(3):432-4, 436.
We describe a simple process for extraction of DNA from amber-entombed fossils and museum specimens that is suitable for enzymatic amplification by PCR. Five to ten milligrams of the macerated specimen were mixed in 300 microliters of silica matrix and shaken at 55 degrees C for 1 h in a sterile, screw-capped microcentrifuge tube. After incubation, the silica matrix was transferred to the upper chamber of a SpinFilter, centrifuged at maximum speed for 1 min and then washed twice with 500 microliters of wash solution and the DNA eluted with 50 microliters of TE buffer. The eluate was used as template for PCR, and the results were evaluated by electrophoresis and nucleotide sequence analysis. All samples tested yielded positive results, which were subsequently verified by sequence analysis. It appears, at least in our hands, that the procedure described here is a rapid and efficient way of obtaining small amounts of DNA for PCR in museum and fossilized specimens.
我们描述了一种从琥珀包裹的化石和博物馆标本中提取DNA的简单方法，该方法适用于通过PCR进行酶促扩增。将5至10毫克研磨后的标本与300微升硅胶基质混合，在无菌带螺旋盖的微量离心管中于55摄氏度振荡1小时。孵育后，将硅胶基质转移至SpinFilter的上腔室，以最大速度离心1分钟，然后用500微升洗涤液洗涤两次，并用50微升TE缓冲液洗脱DNA。洗脱液用作PCR模板，并通过电泳和核苷酸序列分析评估结果。所有测试样品均产生阳性结果，随后通过序列分析进行了验证。至少在我们手中，这里描述的方法似乎是一种从博物馆标本和化石标本中快速有效地获取少量用于PCR的DNA的方法。
