Mutation Spectrum of the Survival of Motor Neuron 1 and Functional Analysis of Variants in Chinese Spinal Muscular Atrophy Yu-jin Qu, Jin-li Bai, Yan-yan.

Slides:

Advertisements

Similar presentations

Figure 1. RT–PCR identification of an abnormal transcript of the PTPN6 gene in normal and leukemic bone marrow cells and cell line. (a) Diagrammatic representation.

Advertisements

From: Three Novel Pax6 Alleles in the Mouse Leading to the Same Small-Eye Phenotype Caused by Different Consequences at Target Promoters Invest. Ophthalmol.

Performance Evaluation of the TheraTyper-GJB2 Assay for Detection of GJB2 Gene Mutations Ji-Yong Chun, Soo-Kyung Shin, Kyung Tae Min, Woojae Cho, Jaeil.

Epidermolysis Bullosa: Novel and De Novo Premature Termination Codon and Deletion Mutations in the Plectin Gene Predict Late-Onset Muscular Dystrophy

Novel Functional Single Nucleotide Polymorphisms in the Latent Transforming Growth Factor-β Binding Protein-1L Promoter Tomomi Higashi, Satoru Kyo, Masaki.

Simultaneous Genotyping of α-Thalassemia Deletional and Nondeletional Mutations by Real-Time PCR–Based Multicolor Melting Curve Analysis Qiuying Huang,

Crucial Roles of MZF1 and Sp1 in the Transcriptional Regulation of the Peptidylarginine Deiminase Type I Gene (PADI1) in Human Keratinocytes Sijun Dong,

Genotyping of Chimerical BCR-ABL1 RNA in Chronic Myeloid Leukemia by Integrated DNA Chip Jong-Hun Kang, Hyun-Gyung Goh, Sang-Ho Chae, Sung-Yong Kim,

Betaine, Dimethyl Sulfoxide, and 7-Deaza-dGTP, a Powerful Mixture for Amplification of GC-Rich DNA Sequences Marco Musso, Renata Bocciardi, Sara Parodi,

Yanggu Shi, Sharon F. Terry, Patrick F. Terry, Lionel G

Application of Single-Molecule Amplification and Resequencing Technology for Broad Surveillance of Plasma Mutations in Patients with Advanced Lung Adenocarcinoma

Application of Single-Molecule Amplification and Resequencing Technology for Broad Surveillance of Plasma Mutations in Patients with Advanced Lung Adenocarcinoma

Philippe Szankasi, Mohamed Jama, David W. Bahler

Herlitz Junctional Epidermolysis Bullosa: Novel and Recurrent Mutations in the LAMB3 Gene and the Population Carrier Frequency Aoi Nakano, Ellen Pfendner,

Comparison of High-Resolution Melting Analysis, TaqMan Allelic Discrimination Assay, and Sanger Sequencing for Clopidogrel Efficacy Genotyping in Routine.

A Locked Nucleic Acid Clamp-Mediated PCR Assay for Detection of a p53 Codon 249 Hotspot Mutation in Urine Selena Y. Lin, Veerpal Dhillon, Surbhi Jain,

A Clinical Grade Sequencing-Based Assay for CEBPA Mutation Testing

Eija Siintola, Meral Topcu, Nina Aula, Hannes Lohi, Berge A

Hou-Sung Jung, Gregory J. Tsongalis, Joel A. Lefferts

Quantitative Analysis of Survival Motor Neuron Copies: Identification of Subtle SMN1 Mutations in Patients with Spinal Muscular Atrophy, Genotype-Phenotype.

Undifferentiated Small Round Cell Sarcomas with Rare EWS Gene Fusions

Simultaneous Genotyping of α-Thalassemia Deletional and Nondeletional Mutations by Real-Time PCR–Based Multicolor Melting Curve Analysis Qiuying Huang,

A Novel DHPLC-Based Procedure for the Analysis of COL1A1 and COL1A2 Mutations in Osteogenesis Imperfecta Antonella Fuccio, Mariangela Iorio, Felice Amato,

Thomas W. Prior, Scott J. Bridgeman

A Novel Alu-Like Element Rearranged in the Dystrophin Gene Causes a Splicing Mutation in a Family with X-Linked Dilated Cardiomyopathy Alessandra Ferlini,

Performance Evaluation of the TheraTyper-GJB2 Assay for Detection of GJB2 Gene Mutations Ji-Yong Chun, Soo-Kyung Shin, Kyung Tae Min, Woojae Cho, Jaeil.

Toward a Survey of Somatic Mutation of the NF1 Gene in Benign Neurofibromas of Patients with Neurofibromatosis Type 1 Ingrid Eisenbarth, Kim Beyer, Winfrid.

Andrea Gaedigk, Amanda K. Riffel, J. Steven Leeder

Volume 54, Issue 3, Pages (September 1998)

Novel Mutations in the LAMC2 Gene in Non-Herlitz Junctional Epidermolysis Bullosa: Effects on Laminin-5 Assembly, Secretion, and Deposition Daniele Castiglia,

Pathogenicity Evaluation of BRCA1 and BRCA2 Unclassified Variants Identified in Portuguese Breast/Ovarian Cancer Families Catarina Santos, Ana Peixoto,

Mutations in a Novel Gene with Transmembrane Domains Underlie Usher Syndrome Type 3 Tarja Joensuu, Riikka Hämäläinen, Bo Yuan, Cheryl Johnson, Saara.

Epidermolysis Bullosa: Novel and De Novo Premature Termination Codon and Deletion Mutations in the Plectin Gene Predict Late-Onset Muscular Dystrophy

Double Heterozygosity for a RET Substitution Interfering with Splicing and an EDNRB Missense Mutation in Hirschsprung Disease Alberto Auricchio, Paola.

Analysis of Rare APC Variants at the mRNA Level

The c.1364C>A (p.A455E) Mutation in the CFTR Pseudogene Results in an Incorrectly Assigned Carrier Status by a Commonly Used Screening Platform Kristin.

Size Polymorphisms in the Human Ultrahigh Sulfur Hair Keratin-Associated Protein 4, KAP4, Gene Family Naoyuki Kariya, Yutaka Shimomura, Masaaki Ito

Volume 38, Issue 1, Pages (April 2010)

Identification and differential expression of human collagenase-3 mRNA species derived from internal deletion, alternative splicing, and different polyadenylation.

Splice Site and Deletion Mutations in Keratin (KRT1 and KRT10) Genes: Unusual Phenotypic Alterations in Scandinavian Patients with Epidermolytic Hyperkeratosis

Structure of the GM2A Gene: Identification of an Exon 2 Nonsense Mutation and a Naturally Occurring Transcript with an In-Frame Deletion of Exon 2 Biao.

A Highly Sensitive Genetic Protocol to Detect NF1 Mutations

Evidence That Translation Reinitiation Leads to a Partially Functional Menkes Protein Containing Two Copper-Binding Sites Marianne Paulsen, Connie Lund,

A Multi-Exonic BRCA1 Deletion Identified in Multiple Families through Single Nucleotide Polymorphism Haplotype Pair Analysis and Gene Amplification with.

Larissa V. Furtado, Helmut C. Weigelin, Kojo S. J

A Homozygous Nonsense Mutation in Type XVII Collagen Gene (COL17A1) Uncovers an Alternatively Spliced mRNA Accounting for an Unusually Mild Form of Non-Herlitz.

Larissa V. Furtado, Helmut C. Weigelin, Kojo S. J

Nonsense mutation of EMX2 is potential causative for uterus didelphysis: first molecular explanation for isolated incomplete müllerian fusion Shan Liu,

Evaluating the Effect of Unclassified Variants Identified in MMR Genes Using Phenotypic Features, Bioinformatics Prediction, and RNA Assays Lucia Pérez-Cabornero,

Amplification Refractory Mutation System, a Highly Sensitive and Simple Polymerase Chain Reaction Assay, for the Detection of JAK2 V617F Mutation in Chronic.

A Mutation in the Variable Repeat Region of the Aggrecan Gene (AGC1) Causes a Form of Spondyloepiphyseal Dysplasia Associated with Severe, Premature.

Novel Polymorphism in the FMR1 Gene Resulting in a “Pseudodeletion” of FMR1 in a Commonly Used Fragile X Assay Thomas M. Daly, Arash Rafii, Rick A. Martin,

Sex-Linked period Genes in the Silkmoth, Antheraea pernyi

Feras M. Hantash, Arlene Rebuyon, Mei Peng, Joy B

Volume 58, Issue 2, Pages (August 2000)

Relationship between CFTR and CTRC Variants and the Clinical Phenotype in Late- Onset Cystic Fibrosis Disease with Chronic Pancreatitis Anna C. Tomaiuolo,

Optimized Allele-Specific Real-Time PCR Assays for the Detection of Common Mutations in KRAS and BRAF Alois H. Lang, Heinz Drexel, Simone Geller-Rhomberg,

Assessing the Functional Characteristics of Synonymous and Nonsynonymous Mutation Candidates by Use of Large DNA Constructs A.M. Eeds, D. Mortlock, R.

Mutation Analysis of the Entire PKD1 Gene: Genetic and Diagnostic Implications Sandro Rossetti, Lana Strmecki, Vicki Gamble, Sarah Burton, Vicky Sneddon,

Maple Syrup Urine Disease: Identification and Carrier-Frequency Determination of a Novel Founder Mutation in the Ashkenazi Jewish Population Lisa Edelmann,

Wook Lew Journal of Investigative Dermatology

Establishment of a Molecular Diagnostic System for Spinal Muscular Atrophy Jian Zeng, Yanhong Lin, Aizhen Yan, Longfeng Ke, Zhongyong Zhu, Fenghua Lan

Bart A. Jessen, Marjorie A. Phillips, Robert H. Rice

Two Exon-Skipping Mutations as the Molecular Basis of Succinic Semialdehyde Dehydrogenase Deficiency (4-Hydroxybutyric Aciduria) Ken L. Chambliss, Debra.

Intragenic telSMN Mutations: Frequency, Distribution, Evidence of a Founder Effect, and Modification of the Spinal Muscular Atrophy Phenotype by cenSMN.

Mutation of the Ca2+ Channel β Subunit Gene Cchb4 Is Associated with Ataxia and Seizures in the Lethargic (lh) Mouse Daniel L Burgess, Julie M Jones,

Quantitative Analysis of Survival Motor Neuron Copies: Identification of Subtle SMN1 Mutations in Patients with Spinal Muscular Atrophy, Genotype-Phenotype.

Exon Skipping in IVD RNA Processing in Isovaleric Acidemia Caused by Point Mutations in the Coding Region of the IVD Gene Jerry Vockley, Peter K. Rogan,

Fang Wang, Yunfeng Wang, Jie Ding, Jiyun Yang Kidney International

Identification of Novel pro-α2(IX) Collagen Gene Mutations in Two Families with Distinctive Oligo-Epiphyseal Forms of Multiple Epiphyseal Dysplasia Paul.

Presentation transcript:

Mutation Spectrum of the Survival of Motor Neuron 1 and Functional Analysis of Variants in Chinese Spinal Muscular Atrophy Yu-jin Qu, Jin-li Bai, Yan-yan Cao, Hong Wang, Yu-wei Jin, Juan Du, Xiu-shan Ge, Wen-hui Zhang, Yan Li, Sheng-xi He, Fang Song The Journal of Molecular Diagnostics Volume 18, Issue 5, Pages 741-752 (September 2016) DOI: 10.1016/j.jmoldx.2016.05.004 Copyright © 2016 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 1 Pattern diagrams illustrating the partial conversions and partial deletions in SMN1. A: Illustration of the different variations associated with SMN1/SMN2 hybrid genes. B: The types of partial deletions in the SMN1 gene. The exons outlined with dotted lines indicate deletions. del, deletion; E, exon. The Journal of Molecular Diagnostics 2016 18, 741-752DOI: (10.1016/j.jmoldx.2016.05.004) Copyright © 2016 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 2 Sequence analysis of novel SMN1 variants. Variants c.-7_9del (p.0) and c.835-5T>G were identified both in g.DNA sequencing and in c.DNA. mRNA samples from patients exhibiting p.Tyr109Cys, p.Ile249Tyrfs*16, or p.Tyr272Trpfs*35 variants were not obtained; these variants were identified using g.DNA sequencing and PCR restriction fragment-length polymorphism or allele-specific PCR (data not shown). The sequencing of variant p.Ile249Tyrfs*16 was performed using the reverse primer. c.DNA, cDNA clone sequencing; g.DNA, genomic DNA sequencing. The Journal of Molecular Diagnostics 2016 18, 741-752DOI: (10.1016/j.jmoldx.2016.05.004) Copyright © 2016 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 3 Level of FL-SMN1 transcript in patients with SMN1 variants. The patients with subtle variants were divided into two groups according to the time of their joining the study. Transcript levels in all cases were detected and compared with those of normal controls and healthy carriers. The normal controls and healthy carrier controls were different between these two groups (normal 1 and normal 2, carrier1 and carrier2). The detected values of the second groups were labeled with yellow background. The FL-SMN1 transcript levels were measured as the number of molecules per nanogram total RNA by absolute real-time PCR as previously described.14,25 The FL-SMN1 transcript levels in the patients with the underlined variants were significantly decreased compared with healthy carriers carrying only one copy of SMN1. ∗Significant difference compared with the carrier controls (P < 0.05). FL, full-length. The Journal of Molecular Diagnostics 2016 18, 741-752DOI: (10.1016/j.jmoldx.2016.05.004) Copyright © 2016 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 4 Restriction digest of SMN transcripts and ex vivo splicing of the c.835-5T>G variant. A: Transcripts for FL-SMN1, FL-SMN2, and Δ7-SMN2 from the normal controls were separated by 6% polyacrylamide gel electrophoresis after digestion with DdeI. In the patient carrying the c.835-5T>G variant, the FL-SMN1 transcript (1259 bp) was difficult to visualize, but the undigested Δ7-SMN1 fragment (1205 bp, red) was more prominent, which also can be seen in his father. B: Ex vivo splicing of the three variants (c.835-5T>C, c.835-5T>G, and c.835-5T>A). This assay was reported in our previous study.19 Lane 1, wild-type SMN1 control plasmid; lane 2, c.863G>T variant (r.835_*3del, p.Gly279Glufs*5); M, indicates the DNA Marker. Lanes 3 to 5, c.835-5T>C, c.835-5T>G, and c.835-5T>A, respectively; lane 6, splicing-positive control plasmid (c.835-1G>A). Fa, father of this patient who carried this variant (two SMN1 copies); FL, full-length; Mo, mother of this patient who carried only one SMN1 copy; N, normal control; Pa, patient with c.835-5T>G. The Journal of Molecular Diagnostics 2016 18, 741-752DOI: (10.1016/j.jmoldx.2016.05.004) Copyright © 2016 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 5 Functional prediction and in vitro expression of the c.-7_9del variant. A: Results of protein prediction using the ORF Finder software (http://www.ncbi.nlm.nih.gov/gorf/gorf.html) with the SMN1 mRNA sequence (NM000344). The translation of wild-type SMN1 mRNA initiates from 64 to 948 nt, coding 294 amino acid. B: Results of protein prediction for the variant c.-7_9del. Twelve proteins were predicted by the ORF Finder software using the c.-7_9del SMN1 mRNA sequence. All were much smaller than the wild-type SMN protein (only 38 to 116 amino acid). Among these, only one predicted protein (645 to 932 nt, 95 amino acid) retained a portion of the C-terminal sequence of wild-type SMN1 (661 to 948 nt, red box). The sequence of this predicted protein is MPGPRLGPGKP……EGRCSHSLN. C: In vitro expression of the FLAG-SMN1 protein from the wild-type SMN1 plasmid (WT), the p.(Tyr277Cys) mutant plasmid, and the c.-7_9del plasmid. The Western blot analysis results showed that none of the full-length and truncated FLAG-SMN fusion proteins were detected from the c.-7_9del plasmid. The empty plasmid (pFLAG-CMV-6a) was used for the mock transfection group. The Journal of Molecular Diagnostics 2016 18, 741-752DOI: (10.1016/j.jmoldx.2016.05.004) Copyright © 2016 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 6 Schematic representation of SMN1 variants. The 18 variants shown in the upper portion are those identified to date in Chinese patients with SMA. The variants in bold denote novel variants identified in this study; the five mutations shown with underline have also been reported in other populations, and the remaining variants have only been reported in China. The two variants in the black box were the most common variants in the Chinese SMA population. The variants shown below have been reported in other populations and the horizontal arrows indicate the most common variants in the Caucasian populations. SMA, spinal muscular atrophy. The Journal of Molecular Diagnostics 2016 18, 741-752DOI: (10.1016/j.jmoldx.2016.05.004) Copyright © 2016 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions