Gomes Newton C Marcial, Fagbola Olajire, Costa Rodrigo, Rumjanek Norma Gouvea, Buchner Arno, Mendona-Hagler Leda, Smalla Kornelia
Biologische Bundesanstalt für Land- und Forstwirtschaft, Institut für Pflanzenvirologie, Mikrobiologie und biologische Sicherheit, Messeweg 11-12, 38104 Braunschweig, Germany.
Appl Environ Microbiol. 2003 Jul;69(7):3758-66. doi: 10.1128/AEM.69.7.3758-3766.2003.
The fungal population dynamics in soil and in the rhizospheres of two maize cultivars grown in tropical soils were studied by a cultivation-independent analysis of directly extracted DNA to provide baseline data. Soil and rhizosphere samples were taken from six plots 20, 40, and 90 days after planting in two consecutive years. A 1.65-kb fragment of the 18S ribosomal DNA (rDNA) amplified from the total community DNA was analyzed by denaturing gradient gel electrophoresis (DGGE) and by cloning and sequencing. A rhizosphere effect was observed for fungal populations at all stages of plant development. In addition, pronounced changes in the composition of fungal communities during plant growth development were found by DGGE. Similar types of fingerprints were observed in two consecutive growth periods. No major differences were detected in the fungal patterns of the two cultivars. Direct cloning of 18S rDNA fragments amplified from soil or rhizosphere DNA resulted in 75 clones matching 12 dominant DGGE bands. The clones were characterized by their HinfI restriction patterns, and 39 different clones representing each group of restriction patterns were sequenced. The cloning and sequencing approach provided information on the phylogeny of dominant amplifiable fungal populations and allowed us to determine a number of fungal phylotypes that contribute to each of the dominant DGGE bands. Based on the sequence similarity of the 18S rDNA fragment with existing fungal isolates in the database, it was shown that the rhizospheres of young maize plants seemed to select the Ascomycetes order Pleosporales, while different members of the Ascomycetes and basidiomycetic yeast were detected in the rhizospheres of senescent maize plants.
通过对直接提取的DNA进行非培养分析,研究了热带土壤中种植的两个玉米品种的土壤和根际中的真菌种群动态,以提供基线数据。连续两年在种植后20、40和90天从六个地块采集土壤和根际样品。通过变性梯度凝胶电泳(DGGE)以及克隆和测序分析从总群落DNA中扩增出的1.65 kb 18S核糖体DNA(rDNA)片段。在植物发育的所有阶段都观察到真菌种群的根际效应。此外,通过DGGE发现植物生长发育过程中真菌群落组成发生了明显变化。在两个连续的生长时期观察到相似类型的指纹图谱。两个品种的真菌模式未检测到主要差异。从土壤或根际DNA扩增出的18S rDNA片段直接克隆得到75个与12个主要DGGE条带匹配的克隆。通过HinfI限制性酶切图谱对克隆进行表征,并对代表每组限制性酶切图谱的39个不同克隆进行测序。克隆和测序方法提供了关于优势可扩增真菌种群系统发育的信息,并使我们能够确定对每个主要DGGE条带有贡献的一些真菌系统型。基于18S rDNA片段与数据库中现有真菌分离株的序列相似性,结果表明,年轻玉米植株的根际似乎选择了子囊菌纲格孢腔菌目,而在衰老玉米植株的根际中检测到子囊菌和担子菌酵母的不同成员。