Rapid and Robust PCR-Based All-Recombinant Cloning Methodology

Rapid and Robust PCR-Based All-Recombinant Cloning Methodology

We report here a PCR-based cloning methodology that requires no post-PCR modifications such as restriction digestion and phosphorylation of the amplified DNA. The advantage of the present method is that it yields only recombinant clones thus eliminating the need for screening. Two DNA amplification reactions by PCR are performed wherein the first reaction amplifies the gene of interest from a s...

Rapid and reliable cloning of PCR products.

Hetero-stagger cloning: efficient and rapid cloning of PCR products.

A variety of methods have been developed for cloning PCR products, including blunt-end cloning (1), restriction cut back (2), ligation-independent cloning (3), uracil DNA–glycosylase (UDG) treatment of uracil-containing deoxyoligonucleotide primers (4,5) and TA cloning (6–8). Blunt-end cloning of PCR products often requires treatment of PCR products to polish the ends (9). Even with treatments,...

Rapid cloning of PCR-derived RAPD probes.

The random-amplified polymorphic DNA (RAPD) technique (5,7) is one of the most useful methods for species identification and studies on the genetic structure of populations of microand macro-parasites. This method generally provides numerous markers that must be cloned and labeled to be used as probes (1,4). These probes can be used to test the specificity and the polymorphism of the RAPD marke...

Rapid cloning of insect transposon insertion junctions using 'universal' PCR.

Very highly degenerate primers with short specific 3' anchor sequences and 5' adaptors were used in conjunction with nested specific primers to amplify large numbers of unknown insertion junctions of the insect retrotransposon Woot, using genomic DNA as template for the polymerase chain reaction (PCR). This technique, sometimes referred to as universal PCR, is a powerful method for molecular ch...