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Molecular cloning of polymorphic markers on RLGS gel using the spot target cloning method
S Hirotsune1, H Shibata, Y Okazaki
1Gene Bank, RIKEN Tsukuba Life Science Center, Institute of Physical and Chemical Research (RIKEN), Ibaraki, Japan.
Abstract:
A new method for target cloning of DNA fragments corresponding to spots on the two-dimensional restriction landmark genomic scanning (RLGS) profile has been developed (targeted spot cloning). We used a Not I restriction trapper to select target DNA fragments from Not I, Eco RV double digests of genomic DNA. The use of the restriction trapper substantially reduces the background clones that are established from the direct recovery of RLGS spot DNA from the two-dimensional gels. Genomic DNA clones were isolated in this study as mouse genome markers for 58 spot loci that were previously characterized using RLGS spot mapping. This method provides a powerful tool for isolating DNA clones after their identification by RLGS system.