[Construction and analysis of a metagenomic library from Tengchong hot spring soil in Yunnan Province].
By a combination of freezing/thawing/ proteinase K-based method and SDS/high-salt/heating treatment, the mixed environmental genomic DNA was isolated directly from a hot spring soil in Tengchong, Yunnan, China. With this method, The DNA yield was up to 1 - 2 microg/g soil. After purification with the Wizard DNA clean up system (Promega, Madison, Wis), the mixed genomic DNA was partially digested with restriction enzyme Pst I. Digested DNA fragments of 3 - 8 kb were recovered from agrose gel and ligated to the pSK (+) vector. The ligation mixture was transformed into DH10B strain, resulting in the construction of a metagenomic library with about 2.5 x 10(4) clones. Restriction enzyme analysis revealed that the average insert is about 4.6 kb. Some novel sequences were identified via sequencing and gene annotation analysis of 30 clones randomly chosen from this library.
Cai
Ying
Y
Institute of Microbiology, Chinese Academy of Science, Beijing 100080, China. caiying81@163.com
Chen
Xiu-Zhen
XZ
Yang
Ke-Qian
KQ
Huang
Li
L
Dong
Zhi-Yang
ZY
D002681:China; D003001:Cloning, Molecular; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D004262:DNA Restriction Enzymes / Q000378:metabolism; D015698:Genomic Library; D045482:Hot Springs / Q000382:microbiology*; D017422:Sequence Analysis, DNA; D012988:Soil Microbiology; D013696:Temperature
D004247:DNA; D004262:DNA Restriction Enzymes