The introduction of PCR technology to the molecular diagnosis of genetic diseases has increased the speed and range of DNA tests available. Previous analyses of dystrophin gene mutations were time consuming, taking weeks to complete, and used radioisotopic methods. Further developments in DNA amplification and post-amplification techniques have made conventional tube PCR redundant. The rapid me...

Avoiding complementarity between primers when designing a PCR assay constitutes a central rule strongly anchored in the mind of the molecular scientist. 3'-complementarity will extend the primers during PCR elongation using one another as template, consequently disabling further possible involvement in traditional target amplification. However, a 5'-complementarity will leave the primers unchan...

1The Rheumatology Section, Department of Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104; 2The Pediatric Rheumatology Section, The Childrens Hospital of Philadelphia, Philadelphia, Pennsylvania 19104; 3The Biotechnology Center, The Wistar Institute of Anatomy and Biology, Philadelphia, Pennsylvania 19104 DNA amplification utilizing the polymerase chain ...

We present an original application of competitive polymerase chain reaction (PCR) for measuring oncogene amplification in DNA from human tumors by simultaneous PCR amplification of genomic DNA with fixed amounts of an internal standard (competitor DNA). Competitors share the same sequence as the target genes but contain an additional 15- to 20-base-pair insert, which allows resolution of the am...

Laser microdissection (LM) provides a useful method for isolating specific cells or tissues from biological samples. Here, we adapted microdissection protocols to allow high-resolution transcript analysis of different tissues from developing Arabidopsis seed. Sufficient RNA ( approximately 50 ng) was extracted from endosperm tissue for RT-PCR. However, to obtain enough RNA for microarray analys...

A sensitive, labor-saving, and easily automatable nonradioactive procedure named APEX-FCS (amplified probe extension detected by fluorescence correlation spectroscopy) has been established to detect specific in vitro amplification of pathogen genomic sequences. As an example, Mycobacterium tuberculosis genomic DNA was subjected to PCR amplification with the Stoffel fragment of Thermus aquaticus...

Feces-based population genetic studies have become increasingly popular. However, polymerase chain reaction (PCR) amplification rates from fecal material vary depending on the species, populations, loci, and extraction protocols. Here, we assessed the PCR amplification success of three microsatellite markers and a segment of the mitochondrial control region of DNA extracted from field-collected...

We report the design and evaluation of PCR primers 63f and 1387r for amplification of 16S rRNA genes from bacteria. Their specificity and efficacy were tested systematically with a bacterial species and environmental samples. They were found to be more useful for 16S rRNA gene amplification in ecological and systematic studies than PCR amplimers that are currently more generally used.

An internal control of amplification was constructed by recombinant PCR to detect PCR inhibitors. This exogenous DNA was included in the reaction mixture and coamplified with the target gene. This detection was successfully applied to the diagnosis of whooping cough by amplification of a fragment of Bordetella pertussis IS481.