End-It™ DNA End-Repair Kit

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Last modified: 2008-07-15 14:57:15

The End-It™ DNA End-Repair Kit is used to convert DNA containing damaged or incompatible 5'- and/or 3'-protruding ends that result from shearing, restriction endonuclease digestion or PCR amplification to 5'-phosphorylated, blunt-end DNA for fast and efficient blunt-end cloning into plasmid, cosmid, fosmid, BAC or other vectors. The conversion to blunt-end DNA is accomplished by exploiting the 5'→3' polymerase and 5'Æ3' exonuclease activities of T4 DNA Polymerase. T4 Polynucleotide Kinase and ATP are also included in the kit for phosphorylation of the 5'-ends of the blunt-ended DNA for subsequent ligation into a cloning vector. The End-It™ DNA End-Repair Kit is identical to the End-Repair Enzyme Mix provided in the pWEB™ Cosmid Cloning Kit, the pWEB-TNC™ Cosmid Cloning Kit, and the EpiFOS™ Fosmid Library Production Kit and the CopyControl™ Fosmid Library Production Kit.

The End-It™ DNA End-Repair Kit contains reagents for 20 end-repair reactions (repair of up to 100µg of genomic DNA). The end-repaired, 5'-phosphorylated DNA can be efficiently ligated into a blunt-end, dephosphorylated cloning vector using the Fast-Link™ DNA Ligation Kit.

Figure 1. DNA fragments containing any type of ends are rapidly and efficiently converted to 5'-phosphorylated, blunt-ended DNA using the End-It™ DNA End-Repair Kit. The end-repaired DNA is ready for blunt-end ligation into the cloning vector of choice using, for example EPICENTRE®'s Fast-Link™ DNA Ligation Kit.

Last modified: 2008-05-14 16:46:46

Applications

• Prepare sheared, nebulised or restriction enzyme digested genomic DNA for blunt-end cloning into plasmid, cosmid, fosmid or BAC vectors
• Prepare PCR-amplified DNA containing an A-overhang for blunt-end cloning

Benefits

• The repaired DNA is blunt-ended and 5'-phosphorylated for immediate blunt-end ligation into plasmid, cosmid, fosmid or BAC vectors
• High specific activity T4 DNA Polymerase and T4 Polynucleotide Kinase are included for complete conversion of 5'- or 3'-overhang DNA to 5-phosphorylated, blunt-ended DNA
• Less costly and more efficient cloning of PCR product containing A-overhangs

Last modified: 2008-05-14 16:46:46

Protocols for: End-It™ DNA End-Repair Kit

(catalogue number ER0720)

Please note: all protocols off site are the responsibility of the products supplier

Last modified: 2008-05-14 16:46:46