Abstract
A novel microRNA (miRNA) quantification method has been developed using one-step rolling circle-quantitative PCR (RC-qPCR) analysis. Vent (exo-) DNA polymerase is firstly utilized to combine a rolling circle amplification (RCA) and qPCR in one step with high sensitivity and specificity in our RC-qPCR assay. Before performing the RC-qPCR, a padlock probe is ligated only when it is perfectly hybridized with miRNA. This ligation-based miRNA assay is highly specific for mature miRNAs, discriminating among related miRNAs that differ by as little as one nucleotide. It exhibits a dynamic range of seven orders of magnitude with a detection limit of 500 aM, and could be also used for the quantification of other small RNA molecules such as short interfering RNAs (siRNAs).
| Original language | English |
|---|---|
| Journal | Analytica Chimica Acta |
| DOIs | |
| Publication status | Published - 2019 Jan 1 |
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Keywords
- MiR-200a
- MiRNA detection
- One-step amplification
- Rolling circle amplification
- Rolling circle-quantitative PCR (RC-qPCR)
- Vent (exo-) DNA polymerase
ASJC Scopus subject areas
- Analytical Chemistry
- Biochemistry
- Environmental Chemistry
- Spectroscopy
Cite this
Ultrasensitive detection of miRNA via one-step rolling circle-quantitative PCR (RC-qPCR). / Xu, Mingcheng; Ye, Jiawei; Yang, Dan; Abdullah AL-maskri, Abdu Ahmed; Hu, Haihong; Jung, Cheulhee; Cai, Sheng; Zeng, Su.
In: Analytica Chimica Acta, 01.01.2019.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Ultrasensitive detection of miRNA via one-step rolling circle-quantitative PCR (RC-qPCR)
AU - Xu, Mingcheng
AU - Ye, Jiawei
AU - Yang, Dan
AU - Abdullah AL-maskri, Abdu Ahmed
AU - Hu, Haihong
AU - Jung, Cheulhee
AU - Cai, Sheng
AU - Zeng, Su
PY - 2019/1/1
Y1 - 2019/1/1
N2 - A novel microRNA (miRNA) quantification method has been developed using one-step rolling circle-quantitative PCR (RC-qPCR) analysis. Vent (exo-) DNA polymerase is firstly utilized to combine a rolling circle amplification (RCA) and qPCR in one step with high sensitivity and specificity in our RC-qPCR assay. Before performing the RC-qPCR, a padlock probe is ligated only when it is perfectly hybridized with miRNA. This ligation-based miRNA assay is highly specific for mature miRNAs, discriminating among related miRNAs that differ by as little as one nucleotide. It exhibits a dynamic range of seven orders of magnitude with a detection limit of 500 aM, and could be also used for the quantification of other small RNA molecules such as short interfering RNAs (siRNAs).
AB - A novel microRNA (miRNA) quantification method has been developed using one-step rolling circle-quantitative PCR (RC-qPCR) analysis. Vent (exo-) DNA polymerase is firstly utilized to combine a rolling circle amplification (RCA) and qPCR in one step with high sensitivity and specificity in our RC-qPCR assay. Before performing the RC-qPCR, a padlock probe is ligated only when it is perfectly hybridized with miRNA. This ligation-based miRNA assay is highly specific for mature miRNAs, discriminating among related miRNAs that differ by as little as one nucleotide. It exhibits a dynamic range of seven orders of magnitude with a detection limit of 500 aM, and could be also used for the quantification of other small RNA molecules such as short interfering RNAs (siRNAs).
KW - MiR-200a
KW - MiRNA detection
KW - One-step amplification
KW - Rolling circle amplification
KW - Rolling circle-quantitative PCR (RC-qPCR)
KW - Vent (exo-) DNA polymerase
UR - http://www.scopus.com/inward/record.url?scp=85065848277&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85065848277&partnerID=8YFLogxK
U2 - 10.1016/j.aca.2019.05.028
DO - 10.1016/j.aca.2019.05.028
M3 - Article
JO - Analytica Chimica Acta
JF - Analytica Chimica Acta
SN - 0003-2670
ER -