Volume 29 Issue 3
Aug.  2014
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XU Li-Mei, WAN Qiu-Hong, WANG Ding. DNA EXTRACTION FROM FORMALIN -FIXED TISSUES AND GENETIC DIVERSITY OF BAIJI( LIPOTES VEXILLIFER)[J]. ACTA HYDROBIOLOGICA SINICA, 2005, 29(3): 272-278.
Citation: XU Li-Mei, WAN Qiu-Hong, WANG Ding. DNA EXTRACTION FROM FORMALIN -FIXED TISSUES AND GENETIC DIVERSITY OF BAIJI( LIPOTES VEXILLIFER)[J]. ACTA HYDROBIOLOGICA SINICA, 2005, 29(3): 272-278.
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DNA EXTRACTION FROM FORMALIN -FIXED TISSUES AND GENETIC DIVERSITY OF BAIJI( LIPOTES VEXILLIFER)

  • Institute of Hydrobiology, The Chinese Academy of Sciences, Wuhan 430072;
    2. The Key Laboratory of Conservation Genetic and Reproductive Biology for Endangered Wild Animals of the Ministry of Education, Hangzhou 310029;
    3. Graduate School of the Chinese Academy of Sciences, Beijing 100039

  • Received Date: August 01, 2004
  • Rev Recd Date: December 19, 2004
  • Published Date: May 24, 2005
    Graphical Abstract
    • Abstract
  • Archival, formalin-fixed tissue is an invaluable resourceformolecular genetic study,but the utilization of nucleic acid of formalin-fixed tissue is generally problematic1Because of the negative effect of formalin on proteinase K and Taq DNA poly-merase,the cross-linking between proteins and DNA, DNA degradation and other reasons, in order to successfully extract and am-plify DNA from formalin-fixed tissue, formalin removal, intensive protein digestion, DNA purification, and PCR optimization should all be considered1In this study, we report an integrated and modified protocol to efficiently extract and amplify DNA from baiji (Lipotes vexillifer)tissues,which were stored in unbuffered formalin for years1We used three pretreatment methods to remove for-malin: gradual dehydration by alcohol, GTE buffer displacement and critical point drying, by comparison the DNA from samples treated by critical point drying is the best in quality and quantity1Intensive proteinase K treatment including increased proteinase K concentration and prolonged digestion time also greatly enhances quality and quantity of DNA1Because DNA from unbuffered formalin fixed tissue was heavily degraded, a DNA target size of approximately 500bp or smaller was found to be optimal for PCR amplification, in addition nested PCR can greatly increase the positive results of amplification1As a result,DNA of 21 baiji specimens were successfully extracted and partial control region of mitochondrial DNA were amplified and sequenced1All the specimens have identical sequences,which indicates very low genetic diversity of baiji.
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    • References (24)
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