Porteous L A, Armstrong J L
Biotechnology Program, United States Environmental Protection Agency, Corvallis, Oregon 97333.
Curr Microbiol. 1993 Aug;27(2):115-8. doi: 10.1007/BF01570868.
A rapid, simple method is used that yields amplifiable fungal and bacterial DNAs directly from soil. DNA is separated from soil contaminants by electrophoresis in low-melting-temperature agarose and used directly in polymerase chain reaction amplification. Fifty 20-mg samples can be processed in one day. Fragments of 16S and 18S ribosomal RNAs are amplified by polymerase chain reaction with DNA extracted from the soil. Universal primers are used that are capable of amplifying ribosomal DNA from a wide variety of bacteria and fungi. Eubacterial and fungal primers are used that are capable of distinguishing between eubacterial and fungal DNAs. Restriction enzyme digests are performed on amplified DNA fragments from five soil samples.
采用一种快速、简单的方法,可直接从土壤中获得可扩增的真菌和细菌DNA。通过在低熔点琼脂糖中进行电泳,将DNA与土壤污染物分离,并直接用于聚合酶链反应扩增。一天内可处理50个20毫克的样品。用从土壤中提取的DNA通过聚合酶链反应扩增16S和18S核糖体RNA片段。使用通用引物,能够扩增来自多种细菌和真菌的核糖体DNA。使用真细菌和真菌引物,能够区分真细菌和真菌DNA。对来自五个土壤样品的扩增DNA片段进行限制性酶切消化。