DNA amplification fingerprinting of mud carp (Cirrhina molitorella) and fast silver staining of DNA profiles

DNA amplification fingerprinting of genome DNA of wild mud carp(Cirrhina molitorella)were obtained using 20 8-mer primers with high GC content .The DNA amplification fingerprinting technique was established,including primer device,and PCR reaction condition.In the condition,primer concentration was relatively high,and 3.5μmol/L was the maximum.DAF had good repetition using primer pn-16(GCGACCTG);the characteristic DNA amplification profiles of Primer pn-16 were identified in mud carp.DNA amplification fingerprinting of some primers revealed DNA polymorphic.Mean while,fast silver staining of DNA profiles in urea-polyacrylamide gel was applied in DNA amplification fingerprinting.The best silver staining condition parameters was obtained,including the silver concentration and length of AgNO_(3) impregnation,the effects of formaldehyde and thiosulfate concentration,and the imaging development temperature.The concentration of AgNO_(3) 1g/L and impregnation of 30min could get optimal sensitivity.The optimal formaldehyde(37%-40%) concentration was 150μL/L.2mg/L Na_(2)S_(2)O_(3) was optimal concentration for reducing nonspecific background staining.The silver staining technique was also applied to the identification of mud carp by RAPD Primer.The silver staining procedure included fixing,rinsing,impregating,developing and stopping.The method of wet and dry preserved gels was established.