发明名称 |
Recombinant phage and methods |
摘要 |
This disclosure provided methods of cloning a phage genome. Also provided are methods of making a recombinant phage genome. In some embodiments the phage genome is engineered to comprise a heterologous nucleic acid sequence, for example a sequence comprising an open reading frame. In some embodiments the phage genome is cloned in a yeast artificial chromosome. Recombinant phage genomes and recombinant phage are also provided. In some embodiments the methods are high throughput methods such as methods of making a plurality of recombinant phage genomes or recombinant phage. Collections of recombinant phage genomes and recombinant phage are also provided. |
申请公布号 |
US9234227(B2) |
申请公布日期 |
2016.01.12 |
申请号 |
US201213627060 |
申请日期 |
2012.09.26 |
申请人 |
Sample6 Technologies, Inc. |
发明人 |
Lu Timothy Kuan Ta;Koeris Michael Sandor;Chevalier Brett Smith;Holder Jason Wyatt;McKenzie Gregory John;Brownell Daniel Robert |
分类号 |
C12N15/00;C12P19/34;C12N15/10;C12N15/81;C12N15/86 |
主分类号 |
C12N15/00 |
代理机构 |
Cooley, LLP |
代理人 |
Cooley, LLP ;Elrifi Ivor;Pavao Matthew |
主权项 |
1. A method of making a recombinant phage genome, comprising:
(a) selecting a starting phage genome and a eukaryotic vector; (b) selecting a stitching oligonucleotide or oligonucleotide duplex for recombination with the starting phage genome and the eukaryotic vector comprising:
(i) detecting an imperfect oligo repeat at an end of the starting phage genome, and selecting a stitching oligonucleotide comprising a sequence from the ends of the starting phage genome and a sequence flanking the double strand break in the eukaryotic vector;(ii) detecting a perfect oligo repeat at an end of the starting phage genome and selecting a oligonucleotide duplex comprising a sequence homologous to the starting phage genome and a sequence homologous to the eukaryotic vector; (c) introducing the starting phage genome and the eukaryotic vector of step (a), and the stitching oligonucleotide or the oligonucleotide duplex of step (b) into a plurality of eukaryotic vector host cells that are not phage host cells; (d) selecting a vector host cell comprising a recombinant vector comprising an intact starting phage genome as a result of insertion of the starting phage genome into the vector; and (e) inserting a heterologous nucleic acid sequence comprising an open reading frame encoding a screenable marker into the intact starting phage genome to provide a recombinant intact phage genome, wherein the screenable marker is operatively linked to a regulatory control sequence. |
地址 |
Cambridge MA US |