Study On The Soil Microbial Diversity In Ginseng Cropping Land By The Method Of DGGE

Ginseng (Panax ginseng C.A.Mey.) is a jealous and highly cropping plants. Although cultivation has 1,700 years of history, even to lead to weakening of ginseng plant growing,.Output, quality and disease resistance characteristics did not significantly reduce the occurrence of a fundamental change. Over the years, many domestic and foreign scholars from different perspectives Ginseng barrier mechanism was extensive, in-depth research, and Ginseng obstacles due to deterioration of soil physical and chemical properties, nutritional imbalances, beneficial soil microorganisms and enzymes reduced and the other factors. In this paper, ginseng rhizosphere soil and takes three-year-ginseng cropping rhizosphere soil, four-year-ginseng cropping rhizosphere soil, not kind of participation of old ginseng and annual ginseng early farming rhizosphere of different growing season soil samples the use of polymerase chain reaction-denaturing gradient gel electrophoresis analysis (PCR-DGGE) analysis of microbial diversity and research in the microbiological point of view in order to clarify the mechanism of Ginseng obstacles, from the microbiological point of view and then solve the obstacles Ginseng and research and development has laid a theoretical foundation of biological agents.1,Three kinds of microbial genome analysis of DNA extraction methods. Method 1 and Method 2 Although the extraction of total DNA extracted from a larger, but the method of extraction of DNA for a very low concentration; method 2 the existence of low purity of the extracted DNA inhibitors (humic acid, pigment, etc.) components, and PCR amplification see popular band, can not follow-up studies; method can extract the three high-purity DNA, PCR amplification and can be suitable for follow-up study. Method 3 and Method 1, the second major difference is an increase of purification process, manual extraction method proves (SDS) extraction of DNA purity there is not enough to defect, obtained by means of manual extraction of DNA must be purificated, in order to the follow-up study. Kit for DNA extraction of soil with high purity, and is conducive to follow-up PCR, was the best choice.2,Nested polymerase chain reaction (PCR) reaction system and conditions were optimized. The first set of PCR amplification, using bacterial universal primers Primer A and Primer B length amplified 16S rDNA; Second PCR amplification to the first set of PCR products as template, primers with Primer C and Primer D amplification 16S rDNA specific area; reconditioning PCR, the second PCR products were diluted to the original concentration of 1 /10 as a template, primers with Primer C and Primer D, remove the second set of PCR amplification in heterodimers. By agarose gel electrophoresis, can be consistent with requirements of the follow-up experiment with universal ideals. 3,The optimized conditions of denaturing gradient gel electrophoresis (DGGE) parameters were determined. Electrophoresis temperature of 60℃; electrophoresis Stage 60V, 1h; the second phase 100V,18h; concentration gradients 40%-60% range, in the stain about 1h. By agarose gel electrophoresis, can be more general.4,PCR-DGGE was used for studying the triennial and four-year-old and uncropped continuous cropping soil and annual primary soil of ginseng. The results showed that there were great differences for the categories of microbiology or bacteria among with triennial and four-year-old and uncropped continuous cropping soil and annual primary soil of ginseng. The common flora for all the soil samples are band B (Uncultured Brucellaceae bacterium, similarity 98%), band D (Ensifer adhaerens), band F (Acinetobacter sp., similarity 97%), band H (Uncultured alpha proteobacterium clone 651, similarity 97%) band I (Stenotrophomonas maltophilia partial, similarity 82%), band K (Gordonia sp. F867, similarity 85%). The special flora of triennial continuous cropping ginseng soil include band E (Uncultured Comamonadaceae bacterium, similarity 93%), band G (Curvibacter lanceolatus strain JPPB B27 similarity 86%), band J (Uncultured Shewanella sp., similarity 95%). The special flora of four-year-old continuous cropping ginseng soil include band A (Uncultured Brucellaceae bacterium, similarity 98%), band N (Rhodoferax sp. IMCC1723, similarity 97%). The special flora of uncropped continuous cropping ginseng soil include band C (Sinorhizobium sp. B69sp, similarity 97%), band L(Propion ibacterium sp., similarity 92%), band M (Uncultured Raoultella sp., similarity 89%).