Cloning regions of the Drosophila genome by microdissection of polytene chromosome DNA and PCR with nonspecific primer.
A simple and rapid procedure to isolate clones carrying sequences from a specific region of the polytene chromosome of Drosophila is demonstrated. The procedure involves microdissection of the region of interest, amplification of the DNA by PCR using a primer designed to prime the synthesis nonspecifically, labeling of the amplified DNA using the random primer method, and screening of a standard library with the probe to identify and isolate clones carrying sequences homologous to the dissected region. This procedure has the potential to replace the difficult procedure of microcloning, as well as facilitate chromosome walking.
Wesley
C S
CS
Department of Ecology and Evolution, State University of New York, Stony Brook 11794.
Ben
M
M
Kreitman
M
M
Hagag
N
N
Eanes
W F
WF
D000818:Animals; D001483:Base Sequence; D001710:Biotin; D002875:Chromosomes / Q000032:analysis*; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics* / Q000302:isolation & purification; D015342:DNA Probes; D004331:Drosophila melanogaster / Q000235:genetics*; D005784:Gene Amplification; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction
D015342:DNA Probes; D001710:Biotin; D004247:DNA