J Integr Plant Biol. ›› 1997, Vol. 39 ›› Issue (2): -.

• Research Articles •    

Comparison and Improvement of Polymorphism Amplification Methods Used for Detecting DNA Diversity of Plants

Hu Zhi-ang, Yun Rui, Zhong Min, Dong Fu-gui, Wang Hong-xin and Qian Ying-qian   

Abstract: A comparative study on randomly amphfied polymorphic DNA (RAPD) and DNA amplification fingerprinting (DAF) used in detecting DNA diversity of natural populations of liaodong oak( Quercus liaotungensis Koidz. ), peashmb ( Caragana ssp. ) and wild soybean ( Glycine soya (L.)Sieb. et Zucc. ) was conducted. In case of using agarose gel electrophoresis with ethidium bromide (EB) staining which was traditionally used in RAPD, both methods generally showed simple profiles with less than 10 bands. By using urea-polyacrylamide gel electrophoresis (Urea-PAGE) with silver staining which was usually used in DAF, the resulted RAPD profiles, similar to those in DAF ones, became much more resolved and consisted of 20 to 40 bands. Similar profile was shared by RAPD and DAF when primer OPA-02 (TACCGAGCTG) and 7.7a (CGAGCTG) were included respectively. It indicated that both methods may have similar mechanism of amplification. After running agarose gel electrophoresis and EB staining the individual bands were cut off and renm in Urea-PAGE with silver staining. It showed that each EB-stained band consists of several fragments with different length. Gene cloning confirmed that a single EB-stained band in agarose gel did contain many DNA fragments with different molecular weight. Low experimental variabihty has been found in both RAPD and DAF even if Taq DNA polymerase was used. An improved method of RAPD with PAGE and silver staining has been successfitUy used in detection of DNA diversity for natural populations of plants.

Key words: Random amplified polymorphic DNA, DNA amplification fingerprinting, Plant natural population, DNA diversity, Polyacrylamide gel electrophoresis, Silver staining

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