Cloning efficiency

Mutation and cloning efficiency.

Increased cloning efficiency by temperature-cycle ligation.

Molecular cloning is a crucial, and often speed-limiting, step in many standard procedures of molecular biology. Ligation of cohesive DNA ends is normally carried out at 12–16 C to ensure a good balance between enzyme activity and stability of annealed DNA overhangs. Low temperatures generally reduce ligase activity, whereas too high temperatures may reduce cloning efficiencies by melting annea...

Improved efficiency M13 cloning using electroporation.

We compared the efficiency of transformation of recombinant M13 DNA obtained using the traditional calcium chloride method with that obtained by electroporatlon. M13 cloning experiments are often hampered by low yields I.e. insufficient numbers of recombinant plaques are generated. Low concentration of the target DNA, poor ligation due to target DNA impurity and small target fragment size are c...

Enhanced efficiency of cloning FACS-sorted mammalian cells.

ethidium bromide, and nucleic acids are visualized on a UV transilluminator. Typically, about 50–100 ng of supercoiled pUC119-derived plasmid DNA, without any limitation concerning the length of the insert, can be detected using this method (Figure 1). This is sufficient to discriminate between transformants containing inserts of various lengths and the control DNA. Although some chromosomal DN...

High-efficiency cloning system for versatile adaptation of DNA fragments.

Recombinant DNA vectors are fundamental tools in molecular biology and genetic engineering. One of the problems frequently encountered in vector construction is the lack of compatible restriction sites between the vectors and the DNA inserts. The conventional approach to solve this problem is to make the DNA fragments blunt-ended (3,4,12) or to add linkers or adaptors (10,11,14) before ligation...