P element transposition in vitro proceeds by a cut-and-paste mechanism and uses GTP as a cofactor.
We have developed an in vitro reaction system for Drosophila P element transposition. Transposition products were recovered by selection in E. coli, and contained simple P element insertions flanked by 8 bp target site duplications as observed in vivo. Transposition required Mg+2 and partially purified P element transposase. Unlike other DNA rearrangement reactions, P element transposition in vitro used GTP as a cofactor; deoxyGTP, dideoxyGTP, or the nonhydrolyzable GTP analogs GMP-PNP or GMP-PCP were also used. Transposon DNA molecules cleaved at the P element termini were able to transpose, but those lacking 3'-hydroxyl groups were inactive. These biochemical data are consistent with genetic data suggesting that P element transposition occurs via a "cut-and-paste" mechanism.
Kaufman
P D
PD
Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142.
Rio
D C
DC
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004251:DNA Transposable Elements / Q000502:physiology*; D004331:Drosophila melanogaster / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D006160:Guanosine Triphosphate / Q000378:metabolism*; D008274:Magnesium / Q000378:metabolism*; D008969:Molecular Sequence Data; D009713:Nucleotidyltransferases / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D013696:Temperature; D019895:Transposases
D004251:DNA Transposable Elements; D006160:Guanosine Triphosphate; D009713:Nucleotidyltransferases; D019895:Transposases; D008274:Magnesium