Reporter gene expression at single-cell level characterized with real-time RT-PCR, chemiluminescence, fluorescence, and electrochemical imaging.
mRNA from single cells was quantified using real-time RT-PCR after recording the address and reporter protein activity with chemiluminescence, fluorescence, and electrochemical techniques, using luciferase, green fluorescent protein, and secreted alkaline phosphatase. mRNA copy number ranging from below 10(3) to 10(7) in single cells showed a lognormal distribution for both externally introduced reporter genes and internally expressed genes. The fluctuation in the gene expression decreased with the increase of the number of cells picked but did not decrease with the increase of mRNA copy number per cell. We found that the correlation coefficients for mRNA and protein expression in logarithmic plot at single-cell level were much lower than 1.00.
Shiku
Hitoshi
H
Graduate School of Environmental Studies, Tohoku University, Aramaki-Aoba, Sendai, Japan. shiku@bioinfo.che.tohoku.ac.jp
Okazaki
Daisuke
D
Suzuki
Junya
J
Takahashi
Yasufumi
Y
Murata
Tatsuya
T
Akita
Hidetaka
H
Harashima
Hideyoshi
H
Ino
Kosuke
K
Matsue
Tomokazu
T
D000469:Alkaline Phosphatase / Q000032:analysis / Q000235:genetics; D002477:Cells / Q000378:metabolism*; D005453:Fluorescence; D015870:Gene Expression; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D008156:Luciferases / Q000032:analysis / Q000235:genetics; D049449:Luminescence; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction
D012333:RNA, Messenger; D049452:Green Fluorescent Proteins; D008156:Luciferases; D000469:Alkaline Phosphatase