TY - JOUR
T1 - Evaluation of seamless ligation cloning extract preparation methods from an Escherichia coli laboratory strain
AU - Okegawa, Yuki
AU - Motohashi, Ken
N1 - Funding Information:
This work was supported by Japan Society for the Promotion of Science (JSPS) KAKENHI Grant 25650037 (to K.M.).
Publisher Copyright:
© 2015 Elsevier Inc. All rights reserved.
PY - 2015/7/27
Y1 - 2015/7/27
N2 - Seamless ligation cloning extract (SLiCE) is a simple and efficient method for DNA cloning without the use of restriction enzymes. Instead, SLiCE uses homologous recombination activities from Escherichia coli cell lysates. To date, SLiCE preparation has been performed using an expensive commercially available lytic reagent. To expand the utility of the SLiCE method, we evaluated different methods for SLiCE preparation that avoid using this reagent. Consequently, cell extracts prepared with buffers containing Triton X-100, which is a common and low-cost nonionic detergent, exhibited sufficient cloning activity for seamless gene incorporation into a vector.
AB - Seamless ligation cloning extract (SLiCE) is a simple and efficient method for DNA cloning without the use of restriction enzymes. Instead, SLiCE uses homologous recombination activities from Escherichia coli cell lysates. To date, SLiCE preparation has been performed using an expensive commercially available lytic reagent. To expand the utility of the SLiCE method, we evaluated different methods for SLiCE preparation that avoid using this reagent. Consequently, cell extracts prepared with buffers containing Triton X-100, which is a common and low-cost nonionic detergent, exhibited sufficient cloning activity for seamless gene incorporation into a vector.
KW - Detergent extraction
KW - Homologous recombination
KW - Seamless DNA cloning
KW - SLiCE
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U2 - 10.1016/j.ab.2015.06.031
DO - 10.1016/j.ab.2015.06.031
M3 - Article
C2 - 26133399
AN - SCOPUS:84937865066
VL - 486
SP - 51
EP - 53
JO - Analytical Biochemistry
JF - Analytical Biochemistry
SN - 0003-2697
ER -