Abstract
Polymerase chain reaction (PCR) is a very powerful tool for qualitative evaluation of nucleic acids due to its high efficiency and convenience. Together with the reverse transcription (RT) reaction, the PCR method has been widely applied to the quantitative measurement of DNA and RNA messages. Since RT-PCR is much more sensitive than all of the traditional methods for quantification of mRNA, including Northern blot, ribonuclease protection, RNA blot, and solution hybridization assays, it is the method of choice for quantitative analyses of low abundance mRNA messages. However, because of the exponential nature of the PCR amplification, RT-PCR quantitation may be problematic, giving false estimates of the abundance of the target messages. By using the constitutively expressed 'housekeeping' gene cyclophilin as a reference gene to normalize mRNA levels, and by taking data from the exponential phase of the PCR amplification, we have developed a rapid and reliable semi-quantitative measurement of the relative abundance of dopamine transporter (DAT) mRNA. The semi-quantitative PCR method has been applied to illustrate its use for the measurement of DAT mRNA in post mortem human brain.
| Original language | English |
|---|---|
| Pages (from-to) | 132-139 |
| Number of pages | 8 |
| Journal | Brain Research Protocols |
| Volume | 4 |
| Issue number | 2 |
| DOIs | |
| State | Published - Jul 1 1999 |
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Keywords
- Dopamine transporter (DAT)
- Human brain
- Polymerase chain reaction (PCR)
- Reverse transcription (RT)
ASJC Scopus subject areas
- Neuroscience(all)
Cite this
Semi-quantitative reverse-transcriptase polymerase chain reaction : An approach for the measurement of target gene expression in human brain. / Chen, Li; Segal, David M.; Mash, Deborah C.
In: Brain Research Protocols, Vol. 4, No. 2, 01.07.1999, p. 132-139.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Semi-quantitative reverse-transcriptase polymerase chain reaction
T2 - An approach for the measurement of target gene expression in human brain
AU - Chen, Li
AU - Segal, David M.
AU - Mash, Deborah C
PY - 1999/7/1
Y1 - 1999/7/1
N2 - Polymerase chain reaction (PCR) is a very powerful tool for qualitative evaluation of nucleic acids due to its high efficiency and convenience. Together with the reverse transcription (RT) reaction, the PCR method has been widely applied to the quantitative measurement of DNA and RNA messages. Since RT-PCR is much more sensitive than all of the traditional methods for quantification of mRNA, including Northern blot, ribonuclease protection, RNA blot, and solution hybridization assays, it is the method of choice for quantitative analyses of low abundance mRNA messages. However, because of the exponential nature of the PCR amplification, RT-PCR quantitation may be problematic, giving false estimates of the abundance of the target messages. By using the constitutively expressed 'housekeeping' gene cyclophilin as a reference gene to normalize mRNA levels, and by taking data from the exponential phase of the PCR amplification, we have developed a rapid and reliable semi-quantitative measurement of the relative abundance of dopamine transporter (DAT) mRNA. The semi-quantitative PCR method has been applied to illustrate its use for the measurement of DAT mRNA in post mortem human brain.
AB - Polymerase chain reaction (PCR) is a very powerful tool for qualitative evaluation of nucleic acids due to its high efficiency and convenience. Together with the reverse transcription (RT) reaction, the PCR method has been widely applied to the quantitative measurement of DNA and RNA messages. Since RT-PCR is much more sensitive than all of the traditional methods for quantification of mRNA, including Northern blot, ribonuclease protection, RNA blot, and solution hybridization assays, it is the method of choice for quantitative analyses of low abundance mRNA messages. However, because of the exponential nature of the PCR amplification, RT-PCR quantitation may be problematic, giving false estimates of the abundance of the target messages. By using the constitutively expressed 'housekeeping' gene cyclophilin as a reference gene to normalize mRNA levels, and by taking data from the exponential phase of the PCR amplification, we have developed a rapid and reliable semi-quantitative measurement of the relative abundance of dopamine transporter (DAT) mRNA. The semi-quantitative PCR method has been applied to illustrate its use for the measurement of DAT mRNA in post mortem human brain.
KW - Dopamine transporter (DAT)
KW - Human brain
KW - Polymerase chain reaction (PCR)
KW - Reverse transcription (RT)
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UR - http://www.scopus.com/inward/citedby.url?scp=0032821395&partnerID=8YFLogxK
U2 - 10.1016/S1385-299X(99)00009-4
DO - 10.1016/S1385-299X(99)00009-4
M3 - Article
VL - 4
SP - 132
EP - 139
JO - Brain Research Protocols
JF - Brain Research Protocols
SN - 1385-299X
IS - 2
ER -