4771cc78b3025326bb1fbb26e71751c3 pmc_32645888.pdf d44bee7d34898169c2342067039f45839a04ead2 pmc_32645888.pdf c7079f0b0d0e861bd3465d55348c11e9b8f1e39bb749b8b3d4cc62f59eab1c47 pmc_32645888.pdf Title: The Need for Establishing a Universal CTG Sizing Method in Myotonic Dystrophy Type 1 Subject: The number of cytosine-thymine-guanine (CTG) repeats (‘CTG expansion size’) in the 3'untranslated region (UTR) region of the dystrophia myotonica-protein kinase (DMPK) gene is a hallmark of myotonic dystrophy type 1 (DM1), which has been related to age of disease onset and clinical severity. However, accurate determination of CTG expansion size is challenging due to its characteristic instability. We compared five different approaches (heat pulse extension polymerase chain reaction [PCR], long PCR-Southern blot [with three different primers sets—1, 2 and 3] and small pool [SP]-PCR) to estimate CTG expansion size in the progenitor allele as well as the most abundant CTG expansion size, in 15 patients with DM1. Our results indicated variability between the methods (although we found no overall differences between long PCR 1 and 2 and SP-PCR, respectively). While keeping in mind the limited sample size of our patient cohort, SP-PCR appeared as the most suitable technique, with an inverse significant correlation found between CTG expansion size of the progenitor allele, as determined by this method, and age of disease onset (r = -0.734, p = 0.016). Yet, in light of the variability of the results obtained with the different methods, we propose that an international agreement is needed to determine which is the most suitable method for assessing CTG expansion size in DM1. Keywords: CTG expansion size; myotonic dystrophy type 1; long PCR; small pool-PCR; age of disease onset Author: Alfonsina Ballester-Lopez, Ian Linares-Pardo, Emma Koehorst, Judit Núñez-Manchón, Guillem Pintos-Morell, Jaume Coll-Cantí, Miriam Almendrote, Giuseppe Lucente, Andrea Arbex, Jonathan J. Magaña, Nadia M. Murillo-Melo, Alejandro Lucia, Darren G. Monckton, Sarah A. Cumming, Alba Ramos-Fransi, Alicia Martínez-Piñeiro and Gisela Nogales-Gadea Creator: LaTeX with hyperref package Producer: pdfTeX-1.40.18 CreationDate: Tue Jul 7 11:20:23 2020 CEST ModDate: Tue Jul 7 11:20:23 2020 CEST Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 9 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 836191 bytes Optimized: no PDF version: 1.5 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- ZKNBHR+URWPalladioL-Ital Type 1 Custom yes yes no 67 0 TZBGYW+URWPalladioL-Bold Type 1 Custom yes yes no 68 0 FXKWJE+VnURWPalladioL-Bold Type 1 Custom yes yes no 69 0 QTLELQ+Pxbsy Type 1 Builtin yes yes no 70 0 QNLQEI+URWPalladioL-Roma Type 1 Custom yes yes no 71 0 LWJOET+VnURWPalladioL Type 1 Custom yes yes no 72 0 OJJAPG+Pxsy Type 1 Builtin yes yes no 73 0 WDLIVJ+Rpxr Type 1 Builtin yes yes no 74 0 HIZBRM+TimesNewRomanPS-BoldItalicMT Type 1C 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HDMHDP+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 249 0 HDMHEA+PalatinoLinotype CID TrueType Identity-H yes yes yes 250 0 HDMHEB+PalatinoLinotype TrueType WinAnsi yes yes no 251 0 HDMHNO+TimesNewRoman CID TrueType Identity-H yes yes yes 252 0 Jhove (Rel. 1.22.1, 2019-04-17) Date: 2023-05-27 03:50:27 CEST RepresentationInformation: pmc_32645888.pdf ReportingModule: PDF-hul, Rel. 1.12.1 (2019-04-17) LastModified: 2023-03-31 11:43:23 CEST Size: 836191 Format: PDF Version: 1.5 Status: Well-Formed and valid SignatureMatches: PDF-hul MIMEtype: application/pdf PDFMetadata: Objects: 0 FreeObjects: 1 IncrementalUpdates: 0 DocumentCatalog: ViewerPreferences: HideToolbar: false HideMenubar: false HideWindowUI: false FitWindow: true CenterWindow: false DisplayDocTitle: false NonFullScreenPageMode: UseNone Direction: L2R ViewArea: CropBox ViewClip: CropBox PrintArea: CropBox PageClip: CropBox PageLayout: SinglePage PageMode: UseNone Outlines: Item: Title: ぉのぴひはつふっぴどはのぜ〴〩਼㰠⽓ 䝯呯 䐠⡳散瑩潮⸲ Item: Title: きちぴづひどちぬびぜ〴ぜ〰ちのつぜ〴ぜ〰きづぴとはつびぜ〴〩਼㰠⽓ 䝯呯 䐠⡳畢獥捴楯渮㈮ㄩ‾㸊⡜㌷㙜㌷㝜〰いぎぁぜ〴ぜ〰ぅへぴひちっぴどはのぜ〴ぜ〰ちのつぜ〴ぜ〰こふぢなづっぴびぜ〴〩਼㰠⽓ 䝯呯 䐠⡳畢獥捴楯渮㈮㈩‾㸊⡜㌷㙜㌷㝜〰えづちぴぜ〴ぜ〰ぐふぬびづぜ〴ぜ〰ぅへぴづのびどはの〭ぐぃげぜ〴〩਼㰠⽓ 䝯呯 䐠⡳畢獥捴楯渮㈮㌩‾㸊⡜㌷㙜㌷㝜〰がはのでぜ〴ぜ〰ぐぃげ〭こはふぴとづひのぜ〴ぜ〰あぬはぴぜ〴〩਼㰠⽓ 䝯呯 䐠⡳畢獥捴楯渮㈮㐩‾㸊⡜㌷㙜㌷㝜〰こねちぬぬぜ〴ぜ〰ぐははぬ〭ぐぃげぜ〴〩਼㰠⽓ 䝯呯 䐠⡳畢獥捴楯渮㈮㔩‾㸊⡜㌷㙜㌷㝜〰こぴちぴどびぴどっちぬぜ〴ぜ〰ぁのちぬべびどびぜ〴〩਼㰠⽓ 䝯呯 䐠⡳散瑩潮⸳ Children: Item: Title: いぎぁぜ〴ぜ〰ぅへぴひちっぴどはのぜ〴ぜ〰ちのつぜ〴ぜ〰こふぢなづっぴびぜ〴〩਼㰠⽓ 䝯呯 䐠⡳畢獥捴楯渮㈮㈩‾㸊⡜㌷㙜㌷㝜〰えづちぴぜ〴ぜ〰ぐふぬびづぜ〴ぜ〰ぅへぴづのびどはの〭ぐぃげぜ〴〩਼㰠⽓ 䝯呯 䐠⡳畢獥捴楯渮㈮㌩‾㸊⡜㌷㙜㌷㝜〰がはのでぜ〴ぜ〰ぐぃげ〭こはふぴとづひのぜ〴ぜ〰あぬはぴぜ〴〩਼㰠⽓ 䝯呯 䐠⡳畢獥捴楯渮㈮㐩‾㸊⡜㌷㙜㌷㝜〰こねちぬぬぜ〴ぜ〰ぐははぬ〭ぐぃげぜ〴〩਼㰠⽓ 䝯呯 䐠⡳畢獥捴楯渮㈮㔩‾㸊⡜㌷㙜㌷㝜〰こぴちぴどびぴどっちぬぜ〴ぜ〰ぁのちぬべびどびぜ〴〩਼㰠⽓ 䝯呯 䐠⡳散瑩潮⸳ Item: Title: えづちぴぜ〴ぜ〰ぐふぬびづぜ〴ぜ〰ぅへぴづのびどはの〭ぐぃげぜ〴〩਼㰠⽓ 䝯呯 䐠⡳畢獥捴楯渮㈮㌩‾㸊⡜㌷㙜㌷㝜〰がはのでぜ〴ぜ〰ぐぃげ〭こはふぴとづひのぜ〴ぜ〰あぬはぴぜ〴〩਼㰠⽓ 䝯呯 䐠⡳畢獥捴楯渮㈮㐩‾㸊⡜㌷㙜㌷㝜〰こねちぬぬぜ〴ぜ〰ぐははぬ〭ぐぃげぜ〴〩਼㰠⽓ 䝯呯 䐠⡳畢獥捴楯渮㈮㔩‾㸊⡜㌷㙜㌷㝜〰こぴちぴどびぴどっちぬぜ〴ぜ〰ぁのちぬべびどびぜ〴〩਼㰠⽓ 䝯呯 䐠⡳散瑩潮⸳ Item: Title: がはのでぜ〴ぜ〰ぐぃげ〭こはふぴとづひのぜ〴ぜ〰あぬはぴぜ〴〩਼㰠⽓ 䝯呯 䐠⡳畢獥捴楯渮㈮㐩‾㸊⡜㌷㙜㌷㝜〰こねちぬぬぜ〴ぜ〰ぐははぬ〭ぐぃげぜ〴〩਼㰠⽓ 䝯呯 䐠⡳畢獥捴楯渮㈮㔩‾㸊⡜㌷㙜㌷㝜〰こぴちぴどびぴどっちぬぜ〴ぜ〰ぁのちぬべびどびぜ〴〩਼㰠⽓ 䝯呯 䐠⡳散瑩潮⸳ Item: Title: こねちぬぬぜ〴ぜ〰ぐははぬ〭ぐぃげぜ〴〩਼㰠⽓ 䝯呯 䐠⡳畢獥捴楯渮㈮㔩‾㸊⡜㌷㙜㌷㝜〰こぴちぴどびぴどっちぬぜ〴ぜ〰ぁのちぬべびどびぜ〴〩਼㰠⽓ 䝯呯 䐠⡳散瑩潮⸳ Item: Title: こぴちぴどびぴどっちぬぜ〴ぜ〰ぁのちぬべびどびぜ〴〩਼㰠⽓ 䝯呯 䐠⡳散瑩潮⸳ Item: Title: げづびふぬぴびぜ〴〩਼㰠⽓ 䝯呯 䐠⡳散瑩潮⸴ Item: Title: いどびっふびびどはのぜ〴〩਼㰠⽓ 䝯呯 䐠⡳散瑩潮⸵ Item: Title: ぃはのっぬふびどはのびぜ〴〩਼㰠⽓ 䝯呯 䐠⡳散瑩潮⸵ Item: Title: げづてづひづのっづび Filters: FilterPipeline: FlateDecode Fonts: Type1: Font: BaseFont: ZKNBHR+URWPalladioL-Ital FontSubset: true FirstChar: 45 LastChar: 121 FontDescriptor: FontName: ZKNBHR+URWPalladioL-Ital Flags: Symbolic FontBBox: -170, -305, 1010, 941 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: TZBGYW+URWPalladioL-Bold FontSubset: true FirstChar: 2 LastChar: 241 FontDescriptor: FontName: TZBGYW+URWPalladioL-Bold Flags: Symbolic FontBBox: -152, -301, 1000, 935 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: FXKWJE+VnURWPalladioL-Bold FontSubset: true FirstChar: 189 LastChar: 243 FontDescriptor: FontName: FXKWJE+VnURWPalladioL-Bold Flags: Symbolic FontBBox: -152, -301, 1000, 935 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