A rapid and simple PCR-based method for analysis of transgenic fish using a restricted amount of fin tissue.
The protocol described in this paper offers a simple and rapid method for PCR analysis of transgenes using a restricted amount of fin tissue from small-sized transgenic fish. A simple preparation of fin lysate using a buffer containing a low concentration of an ionic detergent, SDS (0.01%), followed by neutralization with a second buffer containing higher concentrations of non-ionic detergents NP40 (2%) and Tween 20 (2%) consistently provides a reliable quantity of high-quality DNA template for PCR amplification of transgenes. Based on this protocol, transgenic fish can be clearly distinguished from non-transgenic fish using PCR in a rapid and reproducible manner. Tedious DNA purifications are avoided while fidelity of amplification and efficient identification of transgenic fish are maintained.
Nam
Yoon Kwon
YK
Fish Genetic Manipulation Laboratory CNR), Department of Aquaculture, Pukyong National University, Pusan 608-737, South Korea. yoonknam@pknu.ac.kr
Park
Ji Eun
JE
Kim
Kyoung Kil
KK
Kim
Dong Soo
DS
D000818:Animals; D030801:Animals, Genetically Modified / Q000235:genetics*; D001483:Base Sequence; D017931:DNA Primers; D005399:Fishes / Q000235:genetics*; D016133:Polymerase Chain Reaction
D017931:DNA Primers