A useful Taq-polymerase-based method for subcloning DNA fragments with 5′ protruding ends

Cloning DNA restriction endonuclease fragments with protruding single-stranded ends.

A new method of in vitro recombination was employed to construct plasmids containing lac promoter fragments 64 bp and 144 bp long. The 64 bp HpaII-HhaI fragment contains the binding site for the catabolite activator protein (CAP). The HpaII-HaeIII 144 bp fragment includes the binding sites for RNA polymerase, the lac repressor and CAP. The method utilizes the ability of T4 DNA polymerase to mak...

DNA sequencing using Taq polymerase.

Three DNA polymerases, namely E.coli DNA polymerase 1 (Klenow), reverse transcriptase and T7 DNA polymerase (sequenase), are commonly used for DNA sequencing by the chain termination method of Sanger and colleagues [1). However, the secondary structure of the DNA template can impede the progress of all three polymerases. I have developed a novel procedure in which the thermostable polymerase of...

A simplified protocol for producing Taq DNA polymerase in biology laboratory

Background: Taq DNA polymerase is a very important enzyme for molecular biological studies such as DNA amplification and DNA sequencing by the PCR. It is a standard enzyme that is used in 90% of molecular biology labs today. The aim of this study was to produce Taq DNA polymerase enzyme in E. coli by a reliable, practical, simple and low cost method. Materials and Methods: In this study, the T...

Taq DNA polymerase extension of internal primers blocks polymerase chain reactions allowing differential amplification of molecules with identical 5' and 3' ends.

Polymerase chain reaction (PCR) methodology (1) has become a routine method for selectively amplifying segments of DNA from a wide variety of sources. Amplification of specific sequences is dependent upon an exact match between the template DNA and the oligonucleotide primers. Mismatches at the 3' terminus lead to greatly reduced amplification, with no detectable product when amplified under th...

Quality control PCR: a method for detecting inhibitors of Taq DNA polymerase.