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海南近海30株抗B16细胞活性放线菌的16S rDNA多样性分析
引用本文:闫莉萍,洪葵,胡申才,刘丽华.海南近海30株抗B16细胞活性放线菌的16S rDNA多样性分析[J].微生物学报,2005,45(2):185-190.
作者姓名:闫莉萍  洪葵  胡申才  刘丽华
作者单位:中国热带农业科学院热带生物技术研究所,热带作物生物技术国家重点实验室,海口,570110
基金项目:“8 63”海洋生物技术青年基金 (2 0 0 2AA62 814 0 ),海南省优秀中青年教师科研与教学奖励基金,海南省自然科学基金 (3 0 2 0 7)~~
摘    要:从海南近海 ,包括文昌红树林、海口红树林以及洋浦港等地采集样品 ,经苯酚、SDS、加热等预处理 ,稀释涂布麦芽汁_酵母膏琼脂 (YE)、淀粉酪素琼脂 (SC)、葡萄糖天冬氨酸琼脂 (GA) ,或者直接将样品稀释涂布加有重铬酸钾的高氏一号琼脂 (Gause)和麦芽汁_酵母膏琼脂等进行平板分离。共获得 35 4株放线菌 ,其中有 76株具有不同程度的抗B16细胞毒活性。比较发现加热预处理法和重铬酸钾选择培养法对于广泛分离筛选抗肿瘤活性放线菌不失为一种快速、简便、行之有效的方法。YE、Gause培养基无论在分离到的放线菌总数 ,还是细胞毒活性菌株的比例上都显示了良好的效果。对 30株具有较强抗B16细胞活性的链霉菌进行了扩增性 16SrDNA限制性酶切片段多样性分析 (16SARDRA) ,表明这 30株链霉菌之间有较大的基因差异性。 0 5 0 6 4 2、0 6 0 386和 0 6 0 5 2 4等 3株菌序列分析进一步证明这 3株菌属于链霉菌属 ,其中菌株 0 5 0 6 4 2与其亲缘关系最近的Streptomycescattleya的相似性仅为 95 % ,因此可能是一个新种。

关 键 词:链霉菌  抗肿瘤活性  16SrDNA  16SARDRA
文章编号:0001-6209(2005)02-0185-06
修稿时间:2004年8月23日

16S rDNA diversity analysis of 30 Streptomycetes isolates displaying significant cytotoxic activity against B16 cell from near-shore sediments of Hainan Island
YAN Li-ping,HONG Kui,HU Shen-cai,LIU Li-hua.16S rDNA diversity analysis of 30 Streptomycetes isolates displaying significant cytotoxic activity against B16 cell from near-shore sediments of Hainan Island[J].Acta Microbiologica Sinica,2005,45(2):185-190.
Authors:YAN Li-ping  HONG Kui  HU Shen-cai  LIU Li-hua
Institution:State Key Laboratory of Tropical Crop Biotechnology, Institute of Tropical Biosciences and Biotechnology, Chinese Academy for Tropical Agricuture Sciences, Haikou 571101, China. lpyan@126.com
Abstract:A total of 354 isolates of actinomycetes, of which 76 were detected cytotoxic activity was isolated from near-shore marine samples collected at Wenchang mangrove, DanZhou harbor and YanPu harbor. Four isolation methods were employed, which are SDS pretreatment, phenol pretreatment, heating pretreatment and potassium dichromate selection culture, and media such as'Yeast extract-Malt extract (YE), Glucose-Asprine (GA), Starch-Casin (SC), Starch-KNO3 (Gause) were used. It was showed that heating pretreatment and potassium dichromate selection culture were more considerable methods for extensive isolation of actinomycetes. Medium YE and Gause showed best results in both the total number of actinomycetes and the number of active isolates against tumor cell B16. The genotypic diversity of 30 strains of Streptomycetes possessing strong cytotoxic activity against B16 cell (ID50 > or =200) was analyzed by 16S ARDRA, which resulted in 17 RFLP types, and indicated relatively rich genotypic diversity among these Streptomycetes. 16S rDNA sequence analysis of three strains, 050642, 060386 and 060524 (ID50 > or = 1200) further confirmed that they all belong to Streptomyces genus and strain 050642 was suggested a novel Streptomyces. Spp with the highest similarity of 95% to Streptomyces cattleya.
Keywords:Streptomycetes  Cytotoxic activity  16S rDNA  16S ARDRA
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