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Red/ET Recombineering

Are you tired of finding restriction sites? AS ONE INTERNATIONAL provides high quality and easy-to-use kits and reagents for cloning, sub-cloning, generating transgenic animals, and manipulating DNA constructs. Particularly, Red/ET Recombineering offers unprecedented freedom in gene cloning. It gives you the powerful tools to clone, subclone, and modify DNA at any chosen position. These Red/ET Kits, Selection Cassettes, and Recombinase Plasmids permit precise engineering of DNA molecules of any size, including very large ones such as BACs or the E. coli chromosomes.

Red/ET Quick & Easy BAC Modification Kit (Cat.# K001)

This kit is designed to modify BACs with the revolutionary Red/ET Recombination technology – a powerful tool for introducing modifications such as insertions and deletions into any type of BAC within 2 weeks. Red/ET Recombination uses in vivo recombination and proofreading activity to minimize unwanted mutations in your construct.a

Step 1: The E. coli strain carrying the BAC, which is to be modified, is transformed with the expression plasmid pRedET.

Step 2: The expression of genes mediating Red/ET is induced by the addition of Larabinose and a temperature shift from 30C to 37C. After induction the cells are prepared for electroporation and the linear Tn5-neo cassette (PCR product) flanked by homology arms is electroporated. Red/ET recombination inserts the functional cassette into the target locus. Only colonies carrying the modified BAC will survive Kanamycin selection on the agar plates.

Step 3: The successful integration of the selection cassette will be monitored by PCR or DNA mini preparation.

Red/ET Counter Selection BAC Modification Kit (Cat. # K002)

This is a new version of counter-selection cassette pRpsL-neo for the Red/ET Recombination system. The kit combines the high Red/ET efficiency plus convenient removal of the Red/ET Recombination protein expression plasmid pRedET after recombination. It also has the following features:

  • Based on streptomycin selection
  • Modify bacterial artificial chromosomes (BACs) within 2-3 weeks by using a counter-selection cassette
  • pRpsL-neo counter-selection cassette shows much higher efficiency than pSacB-neo
  • Compatible with bacterial chromosomes and common ColE1 origin plasmids

Red/ET Kits

Cat # Name
K001 Red/ET Quick & Easy BAC Modification Kit (need license to buy)
K002 Red/ET Counter Selection BAC Modification Kit (need license to buy)
K003 Red/ET BAC Subcloning Kit (need license to buy)
K004 Quick & Easy Red/ET Conditional Knock Out Kit – FRT (need license to buy)
K005 Quick & Easy Red/ET Conditional Knock Out Kit – loxP (need license to buy)
K006 Quick & Easy Red/ET E. coli Gene Deletion Kit (need license to buy)
K008 Direct-cloning-proficient E. coli strain GB05-dir (need license to buy)
K009 Recombineering-proficient E. coli strain GB08-red (need license to buy)

 

Selection Cassettes

Cat # Name
A001 Red/ET Neomycin Selection Cassette, Prokaryotic and Eukaryotic (PGK-gb2-neo)
A002 Red/ET Neomycin Selection Cassette, FRT-flanked, Prokaryotic and Eukaryotic (FRT-PGK-gb2-neo-FRT)
A003 Red/ET Neomycin Selection Cassette, loxP-flanked, Prokaryotic and Eukaryotic (loxP-PGK-gb2-neo-loxP)
A004 Red/ET Neomycin Selection Cassette plus loxP site, FRT flanked, Prokaryotic and Eukaryotic (FRT-PGK-gb2-neo-FRT-loxP)
A005 Red/ET Neomycin Selection Cassette plus loxP site 2nd version, FRT flanked, Prokaryotic and Eukaryotic (loxP-FRT-PGK-gb2-neo-FRT)
A006 Red/ET Chloramphenicol Selection Cassette, FRT-flanked (FRT-cm-FRT)
A007 Red/ET Chloramphenicol Selection Cassette, loxP-flanked (loxP-cm-loxP)
A008 Red/ET Ampicillin Selection Cassette, FRT-flanked (FRT-amp-FRT)
A009 Red/ET Ampicillin Selection Cassette, loxP flanked (loxP-amp-loxP)
A010 Red/ET Hygromycin Selection Cassette, FRT-flanked, Prokaryotic and Eukaryotic (FRT-PGK-gb2-hygro-FRT)
A011 Red/ET Hygromycin Selection Cassette, loxP-flanked, Prokaryotic and Eukaryotic (loxP-PGK-gb2-hygro-loxP)
A012 iCre-FRT-neo-FRT (codon improved Cre) with attached FRT flanked, Pro- and Eukaryotic Neomycin selection cassette
A013 iCreERT2-FRT-neo-FRT (Ligand-inducible iCre with attached FRT flanked, Pro- and Eukaryotic Neomycin Selection Cassette)

 

Recombinase Plasmids

Cat # Name
A104 Red/ET Enhanced Flp Expression Plasmid:707-FLPe, tet-selection
A105 Red/ET Enhanced Flp Expression Plasmid:708-FLPe, cm-selection
A106 Enhanced Flp Expression Plasmid:709-FLPe, amp
A107 Enhanced Flp Expression Plasmid:710-FLPe, km
A112 Red/ET Cre Expression Plasmid:705-Cre, cm-selection
A113 Red/ET Cre Expression Plasmid:706-Cre, tet-selection
A114 Cre expression plasmid with ampicillin resistance marker
A201 Red/ET Enhanced Eukaryotic FLP Expression Plasmid: pCAGGS-FLPe (Academic price, requires MTA)
A202 Red/ET Enhanced Eukaryotic FLP Expression Plasmid: pCAGGS-FLPe (Industry price, requires MTA)
A203 Eukaryotic FLPo Recombinase Expression Plasmid: pCAGGS-FLPo (Academic price, requires limited use license)
A204 Eukaryotic Cre Recombinase Expression Plasmid: pCAGGS-Cre (Academic Price)
A205 Eukaryotic Dre Recombinase Expression Plasmid:CAGGS-Dre (Academic price, requires limited use license)
A301 Arabinose Inducible Prokaryotic Cre Recombinase Expression Plasmid: pSC101-BAD-Cre
A302 Arabinose Inducible Prokaryotic Dre Recombinase Expression Plasmid: pSC101-BAD-Dre (Requires limited use license)
A303 Arabinose-inducible Prokaryotic FLPe Recombinase Expression Plasmid: pSC101-BAD-FLPe (tet)