Jacobsen C S
Department of Ecology and Molecular Biology, Royal Veterinary and Agricultural University, Frederiksberg C, Copenhagen, Denmark.
Appl Environ Microbiol. 1995 Sep;61(9):3347-52. doi: 10.1128/aem.61.9.3347-3352.1995.
A magnetic capture-hybridization PCR technique (MCH-PCR) was developed to eliminate the inhibitory effect of humic acids and other contaminants in PCRs targeting specific soil DNA. A single-stranded DNA probe, which was complementary to an internal part of the target gene, was used to coat magnetic beads. After hybridization in a suspension of soil DNA, magnetic extraction of the beads separated the hybrid DNA from all other soil DNA, humic acids, and other interfering soil components. The MCH was followed by PCR amplification of the specific target DNA. In barley rhizosphere soil, detection of a lux gene inserted in a Pseudomonas fluorescens strain could be demonstrated in nonsterile soil samples (0.5 mg). This corresponded to a detection of fewer than 40 bacterial cells per cm of barley root. The MCH-PCR technique greatly improves the current protocols for PCR detection of specific microorganisms or genes in soil because specific target DNA sequences from very small soil samples can be extracted and determined.
一种磁捕获杂交聚合酶链反应技术（MCH-PCR）被开发出来，以消除腐殖酸和其他污染物对针对特定土壤DNA的聚合酶链反应的抑制作用。一个与目标基因内部部分互补的单链DNA探针被用于包被磁珠。在土壤DNA悬浮液中进行杂交后，通过磁珠提取将杂交DNA与所有其他土壤DNA、腐殖酸和其他干扰性土壤成分分离。磁捕获杂交之后是对特定目标DNA进行聚合酶链反应扩增。在大麦根际土壤中，在非无菌土壤样品（0.5毫克）中能够检测到插入荧光假单胞菌菌株中的lux基因。这相当于每厘米大麦根中检测到少于40个细菌细胞。磁捕获杂交聚合酶链反应技术极大地改进了目前在土壤中对特定微生物或基因进行聚合酶链反应检测的方案，因为可以从非常小的土壤样品中提取和测定特定的目标DNA序列。
