| 摘要 |
A method for the generation of DNA fragmentation library based on a transposition reaction in the presence of a transposon end with an engineered cleaveage site providing facilitated downstream handling of the produced DNA fragments, e.g., in the generation of sequencing templates. Transposon nucleic acids comprising a transposon end sequence and an engineered cleaveage site located in the sequence, e.g., in Mu transposon end sequence, are disclosed. |
| 主权项 |
1. An in vitro method for fragmenting DNA, comprising:
a) forming a plurality of transposon complexes by contacting
(i) a plurality of modified transposon end sequences, with(ii) a plurality of MuA transposase enzymes,wherein the plurality of modified transposon end sequences each comprise top and bottom nucleic acid strands, and having MuA R1 and R2 sequences that bind a MuA transposase enzyme, wherein the top or bottom strand that contains the R1 sequence is modified to include a cleavable site, and wherein the cleavable site is selected from a group consisting of a restriction enzyme recognition sequence, a uracil nucleotide base, a plurality of ribose nucleotides, and a methylated nucleotide; b) contacting the plurality of transposon complexes with a plurality of target DNA molecules; and c) incubating the plurality of transposon complexes and the plurality of target DNA molecules under conditions suitable for transposition of the transposon complexes into the target DNA molecules and for fragmenting the target DNA to produce a plurality of fragmented DNA molecules having both ends joined to the modified transposon end sequence, wherein the modified transposon end sequences that are joined to the fragmented DNA molecules include a cleavable site that is selected from a group consisting of a restriction enzyme recognition sequence, a uracil nucleotide base, a plurality of ribose nucleotides, and a methylated nucleotide. |
