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Multiplex polymerase chain reaction (PCR) with color-tagged module-shuffling primers for comparing gene expression levels in various cells
- Source :
- Nucleic Acids Research. 29:84e-84
- Publication Year :
- 2001
- Publisher :
- Oxford University Press (OUP), 2001.
-
Abstract
- A method based on the multiplex polymerase chain reaction (PCR) and gel electrophoresis for the comparative analysis of gene expression levels was developed. Using the method many cDNA fragments from different sources can be compared simultaneously. Competitive PCR amplification of expressed genes from different sources was performed by using ‘module-shuffling primers’ (MPSs). The MPSs (labeled with different fluorophores) consist of sequence modules of 3 or 4 nt. The modules are arranged in different orders in each primer; therefore, the base sequences of the primers are different but their melting temperatures are identical. The genes expressed in different sources are ligated with tags complementary with the MPSs. Tag-ligated fragments are mixed in one tube and amplified at the same amplification efficiency by the MPSs. Amplified fragments are detected separately by multiple-color gel electrophoresis. This method can detect different amounts of each expressed gene, up to a difference in amounts of 30%, and its detection limit is 0.1 amol per assay.
- Subjects :
- DNA, Complementary
Color
Biology
Nucleic Acid Denaturation
Binding, Competitive
Polymerase Chain Reaction
Sensitivity and Specificity
Cell Line
law.invention
law
Complementary DNA
Gene expression
Multiplex polymerase chain reaction
Genetics
Animals
RNA, Messenger
Gene
NAR Methods Online
Polymerase chain reaction
DNA Primers
Fluorescent Dyes
Gel electrophoresis
Base Sequence
Gene Expression Profiling
Temperature
Reproducibility of Results
Molecular biology
Globins
Molecular Weight
Gene expression profiling
Organ Specificity
Calibration
Thermodynamics
Rabbits
Primer (molecular biology)
Subjects
Details
- ISSN :
- 13624962
- Volume :
- 29
- Database :
- OpenAIRE
- Journal :
- Nucleic Acids Research
- Accession number :
- edsair.doi.dedup.....006a4daf26b377da2eda9637a71ca79d