SCSIO OpenIR  > 中科院海洋生物资源可持续利用重点实验室
Isolation of High Molecular Weight DNA from Marine Sponge Bacteria for BAC Library Construction
Ouyang, YC; Dai, SK; Xie, LW; Kumar, MSR; Sun, W; Sun, HM; Tang, DL; Li, X; lixiang@scsio.ac.cn
2010
Source PublicationMARINE BIOTECHNOLOGY
ISSN1436-2228
Volume12Issue:3Pages:318-325
AbstractMetagenomics is a powerful tool for mining the genetic repositories from environmental microorganisms. Bacteria associated with marine sponges (phylum Porifera) are rich sources of biologically active natural products. However, to date, few compounds are discovered from the sponge metagenomic libraries, and the main reason might be the difficulties in recovery of high molecular weight (HMW) DNA from sponge symbionts to construct large insert libraries. Here, we describe a method to recover HMW bacterial DNA from diverse sponges with high quality for bacterial artificial chromosome (BAC) library construction. Microorganisms concentrated from sponges by differential centrifugation were embedded in agarose plugs to lyse out the HMW DNA for recovery. DNA fragments over 436 kb size were recovered from three different types of sponges, Halichondria sp., Haliclona sp., and Xestospongia sp. To evaluate the recovered DNA quality, the diversity of bacterial DNA comprised in the HMW DNA derived from sponge Halichondria sp. was analyzed, and this HMW DNA sample was also cloned into a shuttle BAC vector between Escherichia coli and Streptomyces sp. The results showed that more than five types of bacterial DNA, i.e., Proteobacteria, Nitrospirae, Cyanobacteria, Planctomycetes, and unidentified bacteria, had been recovered by this method, and an average 100 kb size insert DNA in a constructed BAC library demonstrated that the recovered HMW DNA is suitable for metagenomic library construction.
Department[Li, Xiang] Chinese Acad Sci, Key Lab Marine Bioresources Sustainable Utilizat, Guangdong Key Lab Marine Mat Med LMMM GD, S China Sea Inst Oceanol, Guangzhou 510301, Guangdong, Peoples R China; [Ouyang, Yongchang; Dai, Shikun; Xie, Lianwu; Kumar, M. S. Ravi; Sun, Wei; Sun, Huimin; Li, Xiang] Chinese Acad Sci, LMB, LMMM GD, S China Sea Inst Oceanol, Guangzhou 510301, Guangdong, Peoples R China; [Ouyang, Yongchang; Dai, Shikun; Xie, Lianwu; Kumar, M. S. Ravi; Sun, Wei; Sun, Huimin] Chinese Acad Sci, Grad Univ, Beijing 100049, Peoples R China; [Kumar, M. S. Ravi; Tang, Danling] Chinese Acad Sci, LED, RSMEE, S China Sea Inst Oceanol, Guangzhou 510301, Guangdong, Peoples R China
KeywordDna Recovery Marine Sponge Bacterial Artificial Chromosome Metagenomic Library
Subject AreaBiotechnology & Applied Microbiology ; Marine & Freshwater Biology
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Indexed Bysci
Funding ProjectLMB
WOS IDWOS:000278095400010
Citation statistics
Cited Times:12[WOS]   [WOS Record]     [Related Records in WOS]
Document Type期刊论文
Identifierhttp://ir.scsio.ac.cn/handle/344004/3824
Collection中科院海洋生物资源可持续利用重点实验室
Corresponding Authorlixiang@scsio.ac.cn
Recommended Citation
GB/T 7714
Ouyang, YC,Dai, SK,Xie, LW,et al. Isolation of High Molecular Weight DNA from Marine Sponge Bacteria for BAC Library Construction[J]. MARINE BIOTECHNOLOGY,2010,12(3):318-325.
APA Ouyang, YC.,Dai, SK.,Xie, LW.,Kumar, MSR.,Sun, W.,...&lixiang@scsio.ac.cn.(2010).Isolation of High Molecular Weight DNA from Marine Sponge Bacteria for BAC Library Construction.MARINE BIOTECHNOLOGY,12(3),318-325.
MLA Ouyang, YC,et al."Isolation of High Molecular Weight DNA from Marine Sponge Bacteria for BAC Library Construction".MARINE BIOTECHNOLOGY 12.3(2010):318-325.
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