Enzymes for Molecular Biology |
DNA Endonucleases
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Applications
- Rapid sizing of BAC, fosmid, or cosmid clones.
- Generation of restriction maps of BAC, cosmid, or fosmid clone inserts.1
- Linearization of cos site-containing clones for in vitro packaging.
- Specific cleavage of chromosomal DNA for physical mapping.2
Lambda Terminase is an endonuclease encoded by bacteriophage lambda
that recognizes and cleaves DNA at cos sites, generating 5´ overhangs
of 12 bases in length.3 Since the 12-base cos site
sequence is rare, the sizes of inserts in BAC, fosmid, or cosmid clones
can be rapidly determined.4 EPICENTRE's Lambda Terminase is
highly purified to provide reproducible cleavage with minimum nonspecific
nicking activity. A typical digestion requires only 1 unit of Lambda
Terminase for each microgram of DNA. |
 |
| Figure 1. Digestion of BAC clones with Lambda Terminase
gives one band while digestion with Not I often gives multiple
bands. DNA from six randomly chosen BAC clones were digested with Not I
(Panel 1) or Lambda Terminase (Panel 2). Lane M, Lambda Ladder PFG marker;
lanes 1-6, BAC clones; lane BT, BAC-Tracker™ Supercoiled Ladder. |
Unit Definition: One unit of Lambda Terminase linearizes up to 70%
of 1 µg of the cosmid DNA vector pHC79 in 60 minutes at room temperature
under standard assay conditions.
Storage Buffer: 50% glycerol containing 50 mM Tris-HCl (pH 7.5), 0.1
M NaCl, 0.1 mM EDTA, 1 mM DTT, and 0.1% Triton® X-100.
Reaction Buffer: 500 mM Tris-HCl (pH 8.0) and 50 mM MgCl2.
Quality Control: Lambda Terminase is tested in cos site recognition
and cleavage, and is free of detectable RNase and nonspecific DNA exo- and
endonuclease activity.
References
- Rackwits, H.R. et al. (1985) Gene 40, 259.
- Wang, Y. and Wu, R. (1993) Nucleic Acid Res. 21, 2143.
- Higgins, R. et al. (1988) Cell 54, 765.
- EPICENTRE Forum (2004) 11(4), 22
CopyControl™ BAC Cloning Kits (BamH I, EcoR I, or Hind III)
CopyControl™ pCC1BAC™ Cloning-Ready Vectors (BamH I, EcoR I, or Hind III)
CopyControl™ Fosmid Library Production Kit
CopyControl™ HTP Fosmid Library Production Kit
EpiFOS™ Fosmid Library Production Kit
pWEB-TNC™ Cosmid Cloning Kit
pWEB™ Cosmid Cloning Kit
T4 Polynucleotide Kinase, Cloned
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