High-throughput amplicon-based copy number detection of 11 genes in formalin-fixed paraffin-embedded ovarian tumour samples by MLPA-seq

Olga Kondrashova; Clare J. Love; Sebastian Lunke; Arthur L. Hsu; D. Bowtell; G. Chenevix-Trench; A. Green; P. Webb; A. DeFazio; D. Gertig; N. Traficante; S. Fereday; S. Moore; J. Hung; K. Harrap; T. Sadkowsky; N. Pandeya; M. Malt; A. Mellon; R. Robertson; T. Vanden Bergh; M. Jones; P. Mackenzie; J. Maidens; K. Nattress; Y. E. Chiew; A. Stenlake; H. Sullivan; B. Alexander; P. Ashover; S. Brown; T. Corrish; L. Green; L. Jackman; K. Ferguson; K. Martin; A. Martyn; B. Ranieri; J. White; V. Jayde; P. Mamers; L. Bowes; L. Galletta; D. Giles; J. Hendley; K. Alsop; T. Schmidt; H. Shirley; C. Ball; C. Young; Australian Ovarian Cancer Study (AOCS) Group

doi:10.1371/journal.pone.0143006

High-throughput amplicon-based copy number detection of 11 genes in formalin-fixed paraffin-embedded ovarian tumour samples by MLPA-seq

Olga Kondrashova, Clare J. Love, Sebastian Lunke, Arthur L. Hsu, D. Bowtell, G. Chenevix-Trench, A. Green, P. Webb, A. DeFazio, D. Gertig, N. Traficante, S. Fereday, S. Moore, J. Hung, K. Harrap, T. Sadkowsky, N. Pandeya, M. Malt, A. Mellon, R. RobertsonT. Vanden Bergh, M. Jones, P. Mackenzie, J. Maidens, K. Nattress, Y. E. Chiew, A. Stenlake, H. Sullivan, B. Alexander, P. Ashover, S. Brown, T. Corrish, L. Green, L. Jackman, K. Ferguson, K. Martin, A. Martyn, B. Ranieri, J. White, V. Jayde, P. Mamers, L. Bowes, L. Galletta, D. Giles, J. Hendley, K. Alsop, T. Schmidt, H. Shirley, C. Ball, C. Young, Australian Ovarian Cancer Study (AOCS) Group

Environmental Health Sciences

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Abstract

Whilst next generation sequencing can report point mutations in fixed tissue tumour samples reliably, the accurate determination of copy number is more challenging. The conventional Multiplex Ligation-dependent Probe Amplification (MLPA) assay is an effective tool for measurement of gene dosage, but is restricted to around 50 targets due to size resolution of the MLPA probes. By switching from a size-resolved format, to a sequence-resolved format we developed a scalable, high-throughput, quantitative assay. MLPA-seq is capable of detecting deletions, duplications, and amplifications in as little as 5ng of genomic DNA, including from formalin-fixed paraffin-embedded (FFPE) tumour samples. We show that this method can detect BRCA1, BRCA2, ERBB2 and CCNE1 copy number changes in DNA extracted from snap-frozen and FFPE tumour tissue, with 100% sensitivity and >99.5% specificity.

Original language	English (US)
Article number	e0143006
Journal	PloS one
Volume	10
Issue number	11
DOIs	https://doi.org/10.1371/journal.pone.0143006
State	Published - Nov 1 2015

Bibliographical note

Publisher Copyright:
© 2015 Kondrashovaet al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

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Kondrashova, O., Love, C. J., Lunke, S., Hsu, A. L., Bowtell, D., Chenevix-Trench, G., Green, A., Webb, P., DeFazio, A., Gertig, D., Traficante, N., Fereday, S., Moore, S., Hung, J., Harrap, K., Sadkowsky, T., Pandeya, N., Malt, M., Mellon, A., ... Australian Ovarian Cancer Study (AOCS) Group (2015). High-throughput amplicon-based copy number detection of 11 genes in formalin-fixed paraffin-embedded ovarian tumour samples by MLPA-seq. PloS one, 10(11), Article e0143006. https://doi.org/10.1371/journal.pone.0143006

Kondrashova, O, Love, CJ, Lunke, S, Hsu, AL, Bowtell, D, Chenevix-Trench, G, Green, A, Webb, P, DeFazio, A, Gertig, D, Traficante, N, Fereday, S, Moore, S, Hung, J, Harrap, K, Sadkowsky, T, Pandeya, N, Malt, M, Mellon, A, Robertson, R, Vanden Bergh, T, Jones, M, Mackenzie, P, Maidens, J, Nattress, K, Chiew, YE, Stenlake, A, Sullivan, H, Alexander, B, Ashover, P, Brown, S, Corrish, T, Green, L, Jackman, L, Ferguson, K, Martin, K, Martyn, A, Ranieri, B, White, J, Jayde, V, Mamers, P, Bowes, L, Galletta, L, Giles, D, Hendley, J, Alsop, K, Schmidt, T, Shirley, H, Ball, C, Young, C & Australian Ovarian Cancer Study (AOCS) Group 2015, 'High-throughput amplicon-based copy number detection of 11 genes in formalin-fixed paraffin-embedded ovarian tumour samples by MLPA-seq', PloS one, vol. 10, no. 11, e0143006. https://doi.org/10.1371/journal.pone.0143006

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abstract = "Whilst next generation sequencing can report point mutations in fixed tissue tumour samples reliably, the accurate determination of copy number is more challenging. The conventional Multiplex Ligation-dependent Probe Amplification (MLPA) assay is an effective tool for measurement of gene dosage, but is restricted to around 50 targets due to size resolution of the MLPA probes. By switching from a size-resolved format, to a sequence-resolved format we developed a scalable, high-throughput, quantitative assay. MLPA-seq is capable of detecting deletions, duplications, and amplifications in as little as 5ng of genomic DNA, including from formalin-fixed paraffin-embedded (FFPE) tumour samples. We show that this method can detect BRCA1, BRCA2, ERBB2 and CCNE1 copy number changes in DNA extracted from snap-frozen and FFPE tumour tissue, with 100% sensitivity and >99.5% specificity.",

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AU - Bowtell, D.

AU - Chenevix-Trench, G.

AU - Green, A.

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AU - Malt, M.

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AU - Robertson, R.

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AU - Jones, M.

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AU - Maidens, J.

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AU - Chiew, Y. E.

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AU - Ashover, P.

AU - Brown, S.

AU - Corrish, T.

AU - Green, L.

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AU - Martyn, A.

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AU - White, J.

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High-throughput amplicon-based copy number detection of 11 genes in formalin-fixed paraffin-embedded ovarian tumour samples by MLPA-seq

Abstract

Bibliographical note

UN SDGs

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