Analysis of differential display RT-PCR products using fluorescent primers and GENESCAN software. 1997

K R Luehrsen, and L L Marr, and E van der Knaap, and S Cumberledge
PE Applied Biosystems, Foster City, CA, USA.

Differential display reverse transcription PCR (DDRT-PCR) is a procedure used to identify the induction or repression of gene expression. In most DDRT-PCR protocols, radioisotopes are incorporated during PCR and the cDNA products are detected by autoradiography. This report describes the fluorescent labeling of cDNAs and their detection on automated sequencers from PE Applied Biosystems. A fluorescent tag can be incorporated into the PCR product by using either a labeled primer or a labeled dUTP. The fluorescent signals are analyzed by GENESCAN software. Fluorescent DDRT-PCR increases throughput and obviates the handling of hazardous radioisotopes. A PCR cycling profile, expected to give improved reproducibility, is also described. Because amplified cDNAs can't be recovered from the automated sequencer gel, suggestions are given for the identification and recovery of differentially expressed cDNAs.

UI MeSH Term Description Entries
D004331 Drosophila melanogaster A species of fruit fly frequently used in genetics because of the large size of its chromosomes. D. melanogaster,Drosophila melanogasters,melanogaster, Drosophila
D005456 Fluorescent Dyes Chemicals that emit light after excitation by light. The wave length of the emitted light is usually longer than that of the incident light. Fluorochromes are substances that cause fluorescence in other substances, i.e., dyes used to mark or label other compounds with fluorescent tags. Flourescent Agent,Fluorescent Dye,Fluorescent Probe,Fluorescent Probes,Fluorochrome,Fluorochromes,Fluorogenic Substrates,Fluorescence Agents,Fluorescent Agents,Fluorogenic Substrate,Agents, Fluorescence,Agents, Fluorescent,Dyes, Fluorescent,Probes, Fluorescent,Substrates, Fluorogenic
D005786 Gene Expression Regulation Any of the processes by which nuclear, cytoplasmic, or intercellular factors influence the differential control (induction or repression) of gene action at the level of transcription or translation. Gene Action Regulation,Regulation of Gene Expression,Expression Regulation, Gene,Regulation, Gene Action,Regulation, Gene Expression
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia
D012275 Oryza A genus of grass family (Poaceae) that include several rice species. Oryza sativa,Rice,Rices
D012313 RNA A polynucleotide consisting essentially of chains with a repeating backbone of phosphate and ribose units to which nitrogenous bases are attached. RNA is unique among biological macromolecules in that it can encode genetic information, serve as an abundant structural component of cells, and also possesses catalytic activity. (Rieger et al., Glossary of Genetics: Classical and Molecular, 5th ed) RNA, Non-Polyadenylated,Ribonucleic Acid,Gene Products, RNA,Non-Polyadenylated RNA,Acid, Ribonucleic,Non Polyadenylated RNA,RNA Gene Products,RNA, Non Polyadenylated
D012984 Software Sequential operating programs and data which instruct the functioning of a digital computer. Computer Programs,Computer Software,Open Source Software,Software Engineering,Software Tools,Computer Applications Software,Computer Programs and Programming,Computer Software Applications,Application, Computer Software,Applications Software, Computer,Applications Softwares, Computer,Applications, Computer Software,Computer Applications Softwares,Computer Program,Computer Software Application,Engineering, Software,Open Source Softwares,Program, Computer,Programs, Computer,Software Application, Computer,Software Applications, Computer,Software Tool,Software, Computer,Software, Computer Applications,Software, Open Source,Softwares, Computer Applications,Softwares, Open Source,Source Software, Open,Source Softwares, Open,Tool, Software,Tools, Software
D015203 Reproducibility of Results The statistical reproducibility of measurements (often in a clinical context), including the testing of instrumentation or techniques to obtain reproducible results. The concept includes reproducibility of physiological measurements, which may be used to develop rules to assess probability or prognosis, or response to a stimulus; reproducibility of occurrence of a condition; and reproducibility of experimental results. Reliability and Validity,Reliability of Result,Reproducibility Of Result,Reproducibility of Finding,Validity of Result,Validity of Results,Face Validity,Reliability (Epidemiology),Reliability of Results,Reproducibility of Findings,Test-Retest Reliability,Validity (Epidemiology),Finding Reproducibilities,Finding Reproducibility,Of Result, Reproducibility,Of Results, Reproducibility,Reliabilities, Test-Retest,Reliability, Test-Retest,Result Reliabilities,Result Reliability,Result Validities,Result Validity,Result, Reproducibility Of,Results, Reproducibility Of,Test Retest Reliability,Validity and Reliability,Validity, Face
D016133 Polymerase Chain Reaction In vitro method for producing large amounts of specific DNA or RNA fragments of defined length and sequence from small amounts of short oligonucleotide flanking sequences (primers). The essential steps include thermal denaturation of the double-stranded target molecules, annealing of the primers to their complementary sequences, and extension of the annealed primers by enzymatic synthesis with DNA polymerase. The reaction is efficient, specific, and extremely sensitive. Uses for the reaction include disease diagnosis, detection of difficult-to-isolate pathogens, mutation analysis, genetic testing, DNA sequencing, and analyzing evolutionary relationships. Anchored PCR,Inverse PCR,Nested PCR,PCR,Anchored Polymerase Chain Reaction,Inverse Polymerase Chain Reaction,Nested Polymerase Chain Reaction,PCR, Anchored,PCR, Inverse,PCR, Nested,Polymerase Chain Reactions,Reaction, Polymerase Chain,Reactions, Polymerase Chain
D017931 DNA Primers Short sequences (generally about 10 base pairs) of DNA that are complementary to sequences of messenger RNA and allow reverse transcriptases to start copying the adjacent sequences of mRNA. Primers are used extensively in genetic and molecular biology techniques. DNA Primer,Oligodeoxyribonucleotide Primer,Oligodeoxyribonucleotide Primers,Oligonucleotide Primer,Oligonucleotide Primers,Primer, DNA,Primer, Oligodeoxyribonucleotide,Primer, Oligonucleotide,Primers, DNA,Primers, Oligodeoxyribonucleotide,Primers, Oligonucleotide

Related Publications

K R Luehrsen, and L L Marr, and E van der Knaap, and S Cumberledge
March 1998, BioTechniques,
K R Luehrsen, and L L Marr, and E van der Knaap, and S Cumberledge
February 1998, Nucleic acids research,
K R Luehrsen, and L L Marr, and E van der Knaap, and S Cumberledge
September 1994, FEBS letters,
K R Luehrsen, and L L Marr, and E van der Knaap, and S Cumberledge
February 1991, Nihon rinsho. Japanese journal of clinical medicine,
K R Luehrsen, and L L Marr, and E van der Knaap, and S Cumberledge
January 1999, Methods in molecular biology (Clifton, N.J.),
K R Luehrsen, and L L Marr, and E van der Knaap, and S Cumberledge
January 2008, Methods in molecular biology (Clifton, N.J.),
K R Luehrsen, and L L Marr, and E van der Knaap, and S Cumberledge
August 1997, BioTechniques,
K R Luehrsen, and L L Marr, and E van der Knaap, and S Cumberledge
March 1991, Analytical biochemistry,
K R Luehrsen, and L L Marr, and E van der Knaap, and S Cumberledge
September 2001, BioTechniques,
Copied contents to your clipboard!