Simplified Protocol of Solid-Phase cDNA Libraries for Multiple PCR Amplification

Simplified protocol of solid-phase cDNA libraries for multiple PCR amplification.

PCR amplification of cDNA libraries for cloning and screening.

Screening a cDNA library is a common practice for cloning genes of interest in many laboratories. Success of screening is largely dependent on the abundance of cDNAs representing genes of interest and the availability of suitable probes. The procedure involves several rounds of screening to obtain true-positive clones, which might take a week or longer. Typically, each round of screening requir...

Preimplantation diagnosis by whole-genome amplification, PCR amplification, and solid-phase minisequencing of blastomere DNA.

We have developed a new method for preimplantation diagnosis of inherited diseases. Our procedure for the identification of point mutations in single cells combines whole-genome amplification using 15-mer random primers (primer extension preamplification, PEP) with a single locus-specific PCR amplification, followed by detection of the mutation by solid-phase minisequencing. The procedure was e...

Detection and evaluation of non-recombinants in cDNA libraries by multiple cloning region PCR.

Bacteriophages that are routinely used in cDNA libraries do not require any biological selection for forming plaques. Thus parental non-recombinant phages are always found in variable proportions together with recombinant ones in all cDNA libraries. The presence of non-recombinants in significant proportions dilutes the abundance of rare cDNA species and makes library screening difficult. If th...

Simplified method for the telomeric repeat amplification protocol (TRAP).

Since it was first described by Kim et al. in 1994 (4), the telomeric repeat amplification protocol (TRAP) has been used extensively to detect telomerase activity in a variety of tissues. It is a highly sensitive polymerase chain reaction (PCR)-based assay that has been used to detect telomerase activity in a high proportion of cancers of various origins as well as low levels of activity in cer...