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 coil 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. (C) 2015 Elsevier Inc. All rights reserved.