Please use this identifier to cite or link to this item: https://doi.org/10.1016/j.jmoldx.2016.05.002
Title: Defining the Performance Parameters of a Rapid Screening Tool for FMR1 CGG-Repeat Expansions Based on Direct Triplet-Primed PCR and Melt Curve Analysis
Authors: Rajan-Babu, Indhu-Shree
Lian, Mulias 
Tran, Anh H
Dang, Truong T
Le, Huong T-M
Thanh, Minh N
Lee, Caroline G 
Chong, Samuel S 
Keywords: Science & Technology
Life Sciences & Biomedicine
Pathology
FRAGILE-X-SYNDROME
EXPANDED ALLELES
ASSAY
PREVALENCE
GUIDELINES
RESOLUTION
DISORDERS
DIAGNOSIS
GENE
Issue Date: 1-Sep-2016
Publisher: ELSEVIER SCIENCE INC
Citation: Rajan-Babu, Indhu-Shree, Lian, Mulias, Tran, Anh H, Dang, Truong T, Le, Huong T-M, Thanh, Minh N, Lee, Caroline G, Chong, Samuel S (2016-09-01). Defining the Performance Parameters of a Rapid Screening Tool for FMR1 CGG-Repeat Expansions Based on Direct Triplet-Primed PCR and Melt Curve Analysis. JOURNAL OF MOLECULAR DIAGNOSTICS 18 (5) : 719-730. ScholarBank@NUS Repository. https://doi.org/10.1016/j.jmoldx.2016.05.002
Abstract: Population-based screening for CGG-repeat expansions in the fragile X mental retardation 1 (FMR1) gene that cause fragile X syndrome can now be performed more cost-effectively and simply by combining direct triplet-primed PCR (dTP-PCR) with melting curve analysis (MCA). We have now performed a detailed technical validation to define the operational parameters for achieving robust and reliable performance of the FMR1 dTP-PCR MCA assay. We compared the assay's performance on 2 real-time PCR platforms and determined its analytic sensitivity and specificity. We also assessed the assay's performance on DNA isolated from different sources, the effect of differences in CGG-repeat length and AGG-interruption pattern on melt peak temperature (Tm), and the effect of common substances found in DNA solutions on Tms. The assay performed well in distinguishing normal from expansion-carrying samples. The assay had detection sensitivity down to 1 ng and an analytical specificity beyond 150 ng. In addition to peripheral blood DNA, analysis could also be performed on DNA from saliva, buccal swabs, and dried blood spots. Salt increased Tms, glycogen contamination had minimal effect, whereas AGG interruptions lowered Tms. The FMR1 dTP-PCR MCA screening assay is highly sensitive and specific, performs well using DNA from different sources, and is robust and reproducible when reagent concentrations are maintained across all tested samples.
Source Title: JOURNAL OF MOLECULAR DIAGNOSTICS
URI: https://scholarbank.nus.edu.sg/handle/10635/226891
ISSN: 1525-1578
1943-7811
DOI: 10.1016/j.jmoldx.2016.05.002
Appears in Collections:Staff Publications
Elements

Show full item record
Files in This Item:
File Description SizeFormatAccess SettingsVersion 
ISRajanBabu FMR1 dTPMCA JMD 18.719-30 16.pdfPublished version4.4 MBAdobe PDF

CLOSED

None

SCOPUSTM   
Citations

6
checked on Oct 1, 2022

Page view(s)

35
checked on Sep 29, 2022

Download(s)

1
checked on Sep 29, 2022

Google ScholarTM

Check

Altmetric


Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.