塔里木盆地嗜（耐）盐、碱放线菌的分离及其多样性研究
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摘要
放线菌(Actinomycetes)是一类具有重要价值和多种用途的微生物,广泛存在于不同的自然生态环境中。极端环境中存在的放线菌具有不同于一般放线菌的特殊遗传特性、特殊结构和生理功能,以及特殊的代谢产物。盐碱环境作为极端环境的一种,蕴含着大量的嗜(耐)盐、碱放线菌,是极端放线菌生长的理想场所。本研究从塔里木盆地共采集36份盐碱土样,选取2个代表性盐土样,进行2种复合盐、3种预处理方式和6种培养基的筛选试验;同时选取1个代表性碱土样进行干热处理和5种培养基的筛选试验。结果显示,盐环境土样采用120℃1h干热处理,使用印迹法在甘油-精氨酸琼脂培养基、GW1琼脂培养基和高氏一号培养基上分离嗜盐放线菌为宜;碱环境土样自然风干后,使用倍性稀释法涂布在MM琼脂培养基和改良琼脂培养基上分离嗜碱放线菌为宜。
从塔里木盆地土样中分离出568株嗜盐放线菌和175株嗜碱放线菌。对具有代表性的77株放线菌株进行16SrDNA序列分析,结果显示,嗜盐放线菌分属在6个属,分别是链霉菌属(Streptomyces),糖霉菌属(Glycomyces),拟诺卡氏菌属(Nocardiopsis),放线多孢菌属(Actinopolyspora),链单孢菌属(Streptomonospora)和糖糖孢菌属(Saccharomonospora);嗜碱放线菌分属4个属,分别是链霉菌属(Streptomyces)、拟诺卡氏菌属(Nocardiopsis),糖单孢菌属(Saccharomonospora),链单孢菌属(Streptomonospora)。
对已获得16S rDNA序列的菌株进行PKS基因的筛选,检测结果表明,I型PKS基因的阳性率为34.1%,Ⅱ型PKS的阳性率为32.4%,其中链霉菌属和拟诺卡氏菌属是产生聚酮类化合物最丰富的菌源。这些结果显示本实验分离的放线菌菌株不仅具有种属的多样性,在次级代谢合成相关基因上具有更复杂的多样性。
Actinomycetes is a kind of microbe with high important value and actual use, which widely consist in different natural entironment. Actinomycetes consisting in extreme environment had special genetic characteristics, special structures, physiological functions and special metabolizing productions. Salinized environment, which was one of extreme environments and had a lot of the halophilic and alkaline bacteria, was a perfict place that extreme bacteria growed. 36 salinized soil samples were collected from Tarim Basin, especially the Lop Nur. 2 representative salinized soil samples selected, 2 kinds of complex salt, 3 kinds of pretreatment and 6 medium screening tests. At the same time, a representative alkaline dry heat treatment samples and 5 medium screening tests. The results showed that the soil samples of the salt environment in Tarim Basin after 120℃and 1h soil dry heat treatment were coated with the Western blot to glycerol - arginine agar, halophilic Actinomyces on the GW1 agar medium and Gao medium. After the soil samples of alkaline environment naturally air-dried, alkalophilic actinomycetes were separated by dilution with the times of coating to MM agar and improved agar.
There were 568 halophilic actinomycetes and 175 alkaline actinomycetes isolated from the soil samples in Tarim Basin. And 77 representative sequenced strains were analysised by the 16SrDNA sequence. The results showed that the halophilic bacteria were distributed in 6 genera. They were Streptomyce , Glycomyces , Nocardiopsis , Actinopolyspora , Streptomonospora and Saccharomonospora. The alkaline bacteria were distributed in 4 genera. They were Streptomyces , Nocardiopsis , Saccharomonospora and Streptomonospora .
Sequenced strains with 16SrDNA were selected. Test results showed that the type I PKS gene positive rate was 34.1%,Ⅱ-type PKS positive rate was 32.4%, These results indicated that the experimental strains of isolated actinomycetes not only had species diversity of secondary metabolites but also had more complex diversity in the synthesis of related genes.
从塔里木盆地土样中分离出568株嗜盐放线菌和175株嗜碱放线菌。对具有代表性的77株放线菌株进行16SrDNA序列分析,结果显示,嗜盐放线菌分属在6个属,分别是链霉菌属(Streptomyces),糖霉菌属(Glycomyces),拟诺卡氏菌属(Nocardiopsis),放线多孢菌属(Actinopolyspora),链单孢菌属(Streptomonospora)和糖糖孢菌属(Saccharomonospora);嗜碱放线菌分属4个属,分别是链霉菌属(Streptomyces)、拟诺卡氏菌属(Nocardiopsis),糖单孢菌属(Saccharomonospora),链单孢菌属(Streptomonospora)。
对已获得16S rDNA序列的菌株进行PKS基因的筛选,检测结果表明,I型PKS基因的阳性率为34.1%,Ⅱ型PKS的阳性率为32.4%,其中链霉菌属和拟诺卡氏菌属是产生聚酮类化合物最丰富的菌源。这些结果显示本实验分离的放线菌菌株不仅具有种属的多样性,在次级代谢合成相关基因上具有更复杂的多样性。
Actinomycetes is a kind of microbe with high important value and actual use, which widely consist in different natural entironment. Actinomycetes consisting in extreme environment had special genetic characteristics, special structures, physiological functions and special metabolizing productions. Salinized environment, which was one of extreme environments and had a lot of the halophilic and alkaline bacteria, was a perfict place that extreme bacteria growed. 36 salinized soil samples were collected from Tarim Basin, especially the Lop Nur. 2 representative salinized soil samples selected, 2 kinds of complex salt, 3 kinds of pretreatment and 6 medium screening tests. At the same time, a representative alkaline dry heat treatment samples and 5 medium screening tests. The results showed that the soil samples of the salt environment in Tarim Basin after 120℃and 1h soil dry heat treatment were coated with the Western blot to glycerol - arginine agar, halophilic Actinomyces on the GW1 agar medium and Gao medium. After the soil samples of alkaline environment naturally air-dried, alkalophilic actinomycetes were separated by dilution with the times of coating to MM agar and improved agar.
There were 568 halophilic actinomycetes and 175 alkaline actinomycetes isolated from the soil samples in Tarim Basin. And 77 representative sequenced strains were analysised by the 16SrDNA sequence. The results showed that the halophilic bacteria were distributed in 6 genera. They were Streptomyce , Glycomyces , Nocardiopsis , Actinopolyspora , Streptomonospora and Saccharomonospora. The alkaline bacteria were distributed in 4 genera. They were Streptomyces , Nocardiopsis , Saccharomonospora and Streptomonospora .
Sequenced strains with 16SrDNA were selected. Test results showed that the type I PKS gene positive rate was 34.1%,Ⅱ-type PKS positive rate was 32.4%, These results indicated that the experimental strains of isolated actinomycetes not only had species diversity of secondary metabolites but also had more complex diversity in the synthesis of related genes.
引文
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