Development of Genomic Reference Materials for Cystic Fibrosis Genetic Testing

Pratt, Victoria M.; Casini, Michèle; Bridges, Christina; Buller, Arlene; DiAntonio, Lisa L.; Highsmith, W. Edward; Holtegaard, Leonard M.; Muralidharan, Kasinathan; Rohlfs, Elizabeth M.; Tarleton, Jack; Toji, Lorraine; Barker, Shannon D.; Kalman, Lisa V.

doi:10.2353/jmoldx.2009.080149

Cited by 28 publications

(18 citation statements)

References 22 publications

(12 reference statements)

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“…Samples were chosen from cell lines that had been validated as part of the GeT-RM program 8 or sent to participating CAP/ACMG committee member laboratories to verify the mutations before distribution. Results of the MGL2 and MGL5 surveys were evaluated together and, after 2008, separately.…”

Section: Methodsmentioning

confidence: 99%

Molecular genetic testing for cystic fibrosis: laboratory performance on the College of American Pathologists external proficiency surveys

Lyon

Schrijver

Weck

et al. 2015

Genetics in Medicine

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Section: Methodsmentioning

confidence: 99%

Molecular genetic testing for cystic fibrosis: laboratory performance on the College of American Pathologists external proficiency surveys

Lyon

Schrijver

Weck

et al. 2015

Genetics in Medicine

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“…One possible strategy to facilitate the process is to build consortiums to gather and develop reference materials for public use, as has been initiated for CFTR. 47 Moreover, deposition of the novel variants into a "reference library" should be encouraged. Then, a new method can be evaluated with all the deposited variants available at the time of test development, making cross-comparison of the sensitivity between methods and platforms more readily available.…”

Section: Discussionmentioning

confidence: 99%

Mutation Analysis of SLC26A4 for Pendred Syndrome and Nonsyndromic Hearing Loss by High-Resolution Melting

Chen

Tranebjærg

Rendtorff

et al. 2011

The Journal of Molecular Diagnostics

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Pendred syndrome and DFNB4 (autosomal recessive nonsyndromic congenital deafness, locus 4) are associated with autosomal recessive congenital sensorineural hearing loss and mutations in the SLC26A4 gene. Extensive allelic heterogeneity, however, necessitates analysis of all exons and splice sites to identify mutations for individual patients. Although Sanger sequencing is the gold standard for mutation detection, screening methods supplemented with targeted sequencing can provide a cost-effective alternative. One such method, denaturing high-performance liquid chromatography, was developed for clinical mutation detection in SLC26A4. However, this method inherently cannot distinguish homozygous changes from wild-type sequences. High-resolution melting (HRM), on the other hand, can detect heterozygous and homozygous changes cost-effectively, without any post-PCR modifications. We developed a closed-tube HRM mutation detection method specific for SLC26A4 that can be used in the clinical diagnostic setting. Twenty-eight primer pairs were designed to cover all 21 SLC26A4 exons and splice junction sequences. Using the resulting amplicons, initial HRM analysis detected all 45 variants previously identified by sequencing. Subsequently, a 384-well plate format was designed for up to three patient samples per run. Blinded HRM testing on these plates of patient samples collected over 1 year in a clinical diagnostic laboratory accurately detected all variants identified by sequencing. In conclusion, HRM with targeted sequencing is a reliable, simple, and cost-effective method for SLC26A4 mutation screening and detection. Pendred syndrome and DFNB4 (autosomal recessive nonsyndromic congenital deafness, locus 4) are associated with autosomal recessive congenital sensorineural hearing loss. Whereas Pendred syndrome is thought to be the most common cause of syndromic hearing loss, 1,2 DFNB4 is a nonsyndromic form of hearing loss caused by mutations in the same gene. 3 Clinically, both can be associated with temporal bone abnormalities. 4 In addition, an abnormal perchlorate discharge test result and/or euthyroid goiter can be present in patients with Pendred syndrome but not in those with DFNB4. 3,4 This may be due to allelic heterogeneity because differences in residual activity of the protein products render different phenotypes. 5

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“…For clinical laboratories performing molecular diagnostics, participation in proficiency testing/quality assessment programs (PT/EQA) is essential [4][5][6][7]. Disease-specific or methodological PT/EQA protocols are now available in genetic testing [8][9][10][11] and microbiology [12][13][14].…”

Section: Introductionmentioning

confidence: 99%

SPIDIA-DNA: An External Quality Assessment for the pre-analytical phase of blood samples used for DNA-based analyses

Malentacchi

Pazzagli

Simi

et al. 2013

Clinica Chimica Acta

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a b s t r a c t a r t i c l e i n f oBackground: The EC-funded project SPIDIA is aimed to develop evidence-based quality guidelines for the pre-analytical phase of blood samples used for DNA molecular testing. To this purpose, a survey and a pan-European External Quality Assessment (EQA) were implemented. Methods: SPIDIA facility sent to all the participants the same blood sample to be processed without time or temperature limitation. DNA quality parameters performed at SPIDIA facility included: UV spectrophotometric analysis of DNA purity and yield, PCR interferences study by Kineret software and DNA integrity analysis by pulsed field gel electrophoresis. Results: 197 applications have been collected from 30 European countries. A high variability of DNA fragmentation was observed whereas purity, yield and PCR interferences had a narrow distribution within laboratories. A significant difference between the RNase P single copy gene quantity obtained in the DNA samples extracted with the precipitation-based method respect to those obtained with beads and column-based methods was observed. Conclusions:The results of this study will be the basis for implementing a second pan-European EQA and the results of both EQAs will be pooled and will provide the basis for the implementation of evidence-based guidelines for the pre-analytical phase of DNA analysis of blood samples.

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Development of Genomic Reference Materials for Cystic Fibrosis Genetic Testing

Cited by 28 publications

References 22 publications

Molecular genetic testing for cystic fibrosis: laboratory performance on the College of American Pathologists external proficiency surveys

Molecular genetic testing for cystic fibrosis: laboratory performance on the College of American Pathologists external proficiency surveys

Mutation Analysis of SLC26A4 for Pendred Syndrome and Nonsyndromic Hearing Loss by High-Resolution Melting

SPIDIA-DNA: An External Quality Assessment for the pre-analytical phase of blood samples used for DNA-based analyses

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