Diversity Of Endophytic Microbes From Magnolia Denudate Dest Var. Purourascens Rehd Et Wils In Hanzhong Shaanxi Province

Magnoliaceae of the genus Magnolia Magnolia denudate Dest var. purpurascens Rehd et Wils Jun-yu Chen specialties in china, is the national key protected wild plant. The Magnolia denudate Dest var. purpurascens Rehd et Wils is rare, there are reports in the breeding, plant protection, no studies including bacterial diversity, etc. It were used as experimental material to study composition and diversity use of culture-dependent and culture-independent of endophytic microbes in Magnolia denudate Dest var. purpurascens Rehd et Wils. In recent years, culture-independent mainly used to study diversity of endophyte bacteria, but used to study diversity of endophyte fungi rarely reported. This research in order to compare the comprehensive to study composition and diversity of endophytic microbes in Magnolia denudate Dest var. purpurascens Rehd et Wils, for screening its endophytes to prepare bioactive substances work, etc. To further study on its activity of endophytes metabolites and protection has theoretical significance.⑴This research use of culture-dependent received endophytic fungi of Magnolia denudate Dest var. purpurascens Rehd et Wils, when purification of fungi community DNA were extracted, fungi universal primers ITS1/ITS4 were used for 28 S rRNA sequences amplification after molecular identification. This research received 122 endophytic fungi were isolated, a total of 67 strains of morphologic species. Morphological and molecular identification results showed that endophytic fungi belonged to with 5 phyla 16 species. Including Deuteromycotina about 65.7%, contains(Alternaria sp., Phoma sp., Dothiorella sp., Peyronellaea sp., Marssonia sp., Aschersonia sp.); Ascomycota about 22.4%, contains(Ascomycota, Leptosphaerulina sp., Aspergillus sp., Dothideomycetes sp., Coniothyrium sp.); Basidiomycota about 5.9%, contains(Schizophyllum sp.,Basidiomycete); Zygomycota about 2.9%, contains(Rhizopus sp., Mucor sp.) and Glomeromycota about 2.9%, contains Fungal sp..⑵This research use of culture-dependent received endophytic bacterium of Magnolia denudate Dest var. purpurascens Rehd et Wils, bacterium universal primers 27F/1492 R were used for 16 S rRNA sequences amplification after molecular identification. This research received 52 strains endophytic bacterium belonged to with 4 phyla 18 species. Including Bacteroidetes about 7.7%, contains Sphingobacterium sp.; Firmicutes about 15.4%, contains(Planococcus sp., Staphylococcus sp., Bacillus sp., Paenibacillus sp., Desemzia sp.); Proteobacteria about 11.5%, contains(Acinetobacter sp., Pseudomonas sp., Escherichia sp., Klebsiella sp., Cronobacter sp.); Actinobacteria about 40.4 %, contains(Kocuria sp., Micrococcus sp., Microbacterium sp., Brevibacterium sp., Dietzia sp., Actinobacterium sp., Micrococcaceae) and uncultured bacterium about 17.3% of the total.⑶This research has investigated diversity use of culture-independent of endophytic fungi in Magnolia denudate Dest var. purpurascens Rehd et Wils. When total community DNA were extracted, a pair of fungi PCR primers of ITS1/ITS4 were used for endophytic fungi 28 S rRNA gene amplification and a clone library was constructed for the DNA samples. Positive clones were analyzed colony use of pMD-19 T vector universal primers M13-47/M13-48 for purpose fragment to PCR amplification, sequencing analysis cloning gas-bacilli. The results showed that 107 endophytic fungi positive clones belonged to with 3 types 17 species. Including Ascomycota about 57.55%, contains(Phoma sp., Mycoleptodiscus sp., Alternaria sp., Dothideomycetes, Leptosphaeria sp., Ceramothyrium sp., Paraconiothyrium sp., Hyalodendriella sp., Leptosphaerulina sp., Sclerostagonospora sp., Helminthosporium sp., Lophiostoma sp., Microdiplodia sp., Ascochyta sp., Elsinoaceae sp., Dactylaria sp., Sarcinomyces sp.); Uncultured fungi about 21.69% and Fungal endophyte about 20.75%.⑷This research has investigated diversity use of culture-independent of endophytic bacterium in Magnolia denudate Dest var. purpurascens Rehd et Wils. When total community DNA were extracted, a pair of bacterium PCR primers of 799F/1492 R were used for endophytic bacterium 16 S rRNA gene amplification and a clone library was constructed for the DNA samples. Positive clones were analyzed colony use of pMD-19 T vector universal primers M13-47/M13-48 for purpose fragment to PCR amplification, sequencing analysis cloning gas-bacilli. The results showed that 133 endophytic bacterium positive clones belonged to with 4 types 18 species. Including Firmicutes about 53.38%, contains(Firmicutes, Lysinibacillus sp., Lactococcus sp., Lactobacillus sp., Paenibacillus sp., Bacillus sp.); Proteobacteria about 29.32 %, contains(Pseudomonas sp., Pelomonas sp., Providencia sp., Glacial ice sp., Acinetobacter sp., Enterobacter sp., Shigella sp., Aeromonas sp.); Actinobacteria about 6.77%, contains(Streptomyces sp., Actinobacterium sp., Brevibacterium sp., Pantoea sp.); Uncultured bacterium about 9.77%.The results showed that culture-dependent received endophytic fungi 122 strains, 5 phyla 16 species and endophytic bacterial 52 strains, 4 phyla 18 species; culture-independent received 107 endophytic fungi positive clones belonged to 3 types 17 species and 133 endophytic bacterial positive clones belonged to 4 types 18 species. This research relatively comprehensive study of main county composition and diversity of endophytic mycelium in Magnolia denudate Dest var. purpurascens Rehd et Wils, its diversity is abundant.