Competitive RT-PCR to quantify CFTR mRNA in human endometrium.
Because the down-regulation by progesterone of cystic fibrosis transmembrane conductance regulator (CFTR) expression could be a useful specific marker to define the state of implant receptivity in endometrium, a competitive reverse transcription-polymerase chain reaction (RT-PCR) was developed for quantifying the CFTR mRNA concentration in human endometrial samples. A competitor RNA was constructed with the same sequence as the CFTR sequence except for a 20-nucleotide insertion in the middle. The amplified products were separated by polyacrylamide gel electrophoresis. The ratio of CFTR band areas to competitor band areas provided the basis of quantification. Using this competitive RT-PCR, we measured CFTR mRNA in human endometrial samples taken at different periods of the menstrual cycle, in endometriosis, and in hyperplasia. Results show that the method is suitable for measuring the concentration of CFTR mRNA.
Service de Biochimie, Centre Hospitalier Universitaire, Besançon, France.
A
A
Mularoni
G L
GL
Adessi
F
F
Arbez-Gindre
G
G
Agnani
M
M
Nicollier
D001667:Binding, Competitive; D004396:Coloring Agents; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics*; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D004715:Endometriosis / Q000378:metabolism; D004717:Endometrium / Q000737:chemistry* / Q000473:pathology; D004996:Ethidium; D005260:Female; D006801:Humans; D006965:Hyperplasia; D008597:Menstrual Cycle; D009692:Nucleic Acid Heteroduplexes; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis*; D012194:RNA-Directed DNA Polymerase
C505032:CFTR protein, human; D004396:Coloring Agents; D017931:DNA Primers; D009692:Nucleic Acid Heteroduplexes; D012333:RNA, Messenger; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D012194:RNA-Directed DNA Polymerase; D004996:Ethidium