Medical genetics: advances in brief: Point mutations and polymorphisms in the human dystrophin gene identified in genomic DNA sequences amplified by multiplex PCR

SummaryPoint mutations in factor IX genes of four unrelated Chinese patients with hemophilia B have been identified by direct sequencing of amplified genomic DNA fragments. These four mutations occur in exon 8 of the factor IX gene. A C to T transition at nucleotide 30,863 changes codon 248 from Arg (CGA) to a new Stop codon (TGA), described in a previous family as factor IXMalmo3 (Green P M et al., EMBO J 1989; 8: 1067). A G to A transition, at nucleotide 31,051 changes codon 310 from Trp (TGG) to a nonsense or Stop codon (TGA; factor IXchongqing2)- A G to A transition at nucleotide 31,119 changes codon 333 which is for Arg (CGA) in normal factor IX, to one for Gin (CAA) in the variant previously described as factor IXLondon2 (Tsang T C et al., EMBO J 1988; 7: 3009) in a patient with moderately severe hemophilia B. The fourth patient has a novel C to A transversion at nucleotide 31,290, which corresponds to replacement of codon 390 which is for Ala (GCA) in normal factor IX, to one for Glu (GAA) in a patient with moderately severe hemophilia B (factor IXChongqing3)- DNA sequences of amplified fragments from mothers of three showed both their son’s variant and a normal nucleotide at the appropriate position, indicating that they are carriers. The fourth patient’s (factor IXMalmo3) mother, whose DNA was not evaluable, was most probably a carrier because of her low plasma factor IX levels.

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Protein truncation test: Analysis of two novel point mutations at the carboxy-terminus of the human dystrophin gene associated with mental retardation

Human Mutation ◽

10.1002/humu.1380060205 ◽

1995 ◽

Vol 6 (2) ◽

pp. 126-135 ◽

Cited By ~ 18

Author(s):

Sylvie Tuffery ◽

Uwe Lenk ◽

Roland G. Roberts ◽

Christine Coubes ◽

Jacques Demaille ◽

...

Keyword(s):

Mental Retardation ◽

Point Mutations ◽

Dystrophin Gene ◽

Carboxy Terminus ◽

Protein Truncation Test ◽

Test Analysis ◽

Protein Truncation ◽

Human Dystrophin

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Point mutations at the carboxy terminus of the human dystrophin gene: implications for an association with mental retardation in DMD patients

Human Molecular Genetics ◽

10.1093/hmg/2.11.1877 ◽

1993 ◽

Vol 2 (11) ◽

pp. 1877-1881 ◽

Cited By ~ 73

Author(s):

Uwe Lenk ◽

Rita Hanke ◽

Hannelore Thiele ◽

Astrid Speer

Keyword(s):

Mental Retardation ◽

Point Mutations ◽

Dystrophin Gene ◽

Carboxy Terminus ◽

Human Dystrophin

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Characterization of squalene synthase gene from Gymnema sylvestre R. Br.

Beni-Suef University Journal of Basic and Applied Sciences ◽

10.1186/s43088-020-00094-4 ◽

2021 ◽

Vol 10 (1) ◽

Author(s):

Kuldeepsingh A. Kalariya ◽

Ram Prasnna Meena ◽

Lipi Poojara ◽

Deepa Shahi ◽

Sandip Patel

Keyword(s):

Dna Sequences ◽

Genomic Dna ◽

Competitive Inhibition ◽

Sequence Data ◽

Homology Model ◽

Squalene Synthase ◽

Gymnema Sylvestre ◽

Gardenia Jasminoides ◽

Ramachandran Plots ◽

Flanking Regions

Abstract Background Squalene synthase (SQS) is a rate-limiting enzyme necessary to produce pentacyclic triterpenes in plants. It is an important enzyme producing squalene molecules required to run steroidal and triterpenoid biosynthesis pathways working in competitive inhibition mode. Reports are available on information pertaining to SQS gene in several plants, but detailed information on SQS gene in Gymnema sylvestre R. Br. is not available. G. sylvestre is a priceless rare vine of central eco-region known for its medicinally important triterpenoids. Our work aims to characterize the GS-SQS gene in this high-value medicinal plant. Results Coding DNA sequences (CDS) with 1245 bp length representing GS-SQS gene predicted from transcriptome data in G. sylvestre was used for further characterization. The SWISS protein structure modeled for the GS-SQS amino acid sequence data had MolProbity Score of 1.44 and the Clash Score 3.86. The quality estimates and statistical score of Ramachandran plots analysis indicated that the homology model was reliable. For full-length amplification of the gene, primers designed from flanking regions of CDS encoding GS-SQS were used to get amplification against genomic DNA as template which resulted in approximately 6.2-kb sized single-band product. The sequencing of this product through NGS was carried out generating 2.32 Gb data and 3347 number of scaffolds with N50 value of 457 bp. These scaffolds were compared to identify similarity with other SQS genes as well as the GS-SQSs of the transcriptome. Scaffold_3347 representing the GS-SQS gene harbored two introns of 101 and 164 bp size. Both these intronic regions were validated by primers designed from adjoining outside regions of the introns on the scaffold representing GS-SQS gene. The amplification took place when the template was genomic DNA and failed when the template was cDNA confirmed the presence of two introns in GS-SQS gene in Gymnema sylvestre R. Br. Conclusion This study shows GS-SQS gene was very closely related to Coffea arabica and Gardenia jasminoides and this gene harbored two introns of 101 and 164 bp size.

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Rapid Amplification of Uncharacterized Transposon-tagged DNA Sequences from Genomic DNA

Yeast ◽

10.1002/(sici)1097-0061(19970315)13:3<233::aid-yea88>3.0.co;2-e ◽

1997 ◽

Vol 13 (3) ◽

pp. 233-240 ◽

Cited By ~ 70

Author(s):

KRISTIN T. CHUN ◽

HOWARD J. EDENBERG ◽

MARK R. KELLEY ◽

MARK G. GOEBL

Keyword(s):

Dna Sequences ◽

Genomic Dna ◽

Rapid Amplification

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Molecular Markers for Rapidly Identifying Candidate Genes in Chlamydomonas reinhardtii: ERY1 and ERY2 Encode Chloroplast Ribosomal Proteins

Genetics ◽

10.1093/genetics/164.4.1345 ◽

2003 ◽

Vol 164 (4) ◽

pp. 1345-1353

Author(s):

Amber K Bowers ◽

Jennifer A Keller ◽

Susan K Dutcher

Keyword(s):

Molecular Markers ◽

Chlamydomonas Reinhardtii ◽

Candidate Genes ◽

Splice Site ◽

Genomic Dna ◽

Ribosomal Proteins ◽

Genomic Sequence ◽

Point Mutations ◽

Acceptor Site ◽

Wild Type

Abstract To take advantage of available expressed sequence tags and genomic sequence, we have developed 64 PCR-based molecular markers in Chlamydomonas reinhardtii that map to the 17 linkage groups. These markers will allow the rapid association of a candidate gene sequence with previously identified mutations. As proof of principle, we have identified the genes encoded by the ERY1 and ERY2 loci. Mendelian mutations that confer resistance to erythromycin define three unlinked nuclear loci in C. reinhardtii. Candidate genes ribosomal protein L4 (RPL4) and L22 (RPL22) are tightly linked to the ERY1 locus and ERY2 locus, respectively. Genomic DNA for RPL4 from wild type and five mutant ery1 alleles was amplified and sequenced and three different point mutations were found. Two different glycine residues (G102 and G112) are replaced by aspartic acid and both are in the unstructured region of RPL4 that lines the peptide exit tunnel of the chloroplast ribosome. The other two alleles change a splice site acceptor site. Genomic DNA for RPL22 from wild type and three mutant ery2 alleles was amplified and sequenced and revealed three different point mutations. Two alleles have premature stop codons and one allele changes a splice site acceptor site.

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Transcription of alpha-specific genes in Saccharomyces cerevisiae: DNA sequence requirements for activity of the coregulator alpha 1.

Molecular and Cellular Biology ◽

10.1128/mcb.13.11.6866 ◽

1993 ◽

Vol 13 (11) ◽

pp. 6866-6875 ◽

Cited By ~ 20

Author(s):

D C Hagen ◽

L Bruhn ◽

C A Westby ◽

G F Sprague

Keyword(s):

Saccharomyces Cerevisiae ◽

Binding Site ◽

Dna Sequences ◽

Point Mutations ◽

Transcription Activation ◽

Specific Gene ◽

Binding Assays ◽

Weak Binding

Transcription activation of alpha-specific genes in Saccharomyces cerevisiae is regulated by two proteins, MCM1 and alpha 1, which bind to DNA sequences, called P'Q elements, found upstream of alpha-specific genes. Neither MCM1 nor alpha 1 alone binds efficiently to P'Q elements. Together, however, they bind cooperatively in a manner that requires both the P' sequence, which is a weak binding site for MCM1, and the Q sequence, which has been postulated to be the binding site for alpha 1. We analyzed a collection of point mutations in the P'Q element of the STE3 gene to determine the importance of individual base pairs for alpha-specific gene transcription. Within the 10-bp conserved Q sequence, mutations at only three positions strongly affected transcription activation in vivo. These same mutations did not affect the weak binding to P'Q displayed by MCM1 alone. In vitro DNA binding assays showed a direct correlation between the ability of the mutant sequences to form ternary P'Q-MCM1-alpha 1 complexes and the degree to which transcription was activated in vivo. Thus, the ability of alpha 1 and MCM1 to bind cooperatively to P'Q elements is critical for activation of alpha-specific genes. In all natural alpha-specific genes the Q sequence is adjacent to the degenerate side of P'. To test the significance of this geometry, we created several novel juxtapositions of P, P', and Q sequences. When the Q sequence was opposite the degenerate side, the composite QP' element was inactive as a promoter element in vivo and unable to form stable ternary QP'-MCM1-alpha 1 complexes in vitro. We also found that addition of a Q sequence to a strong MCM1 binding site allows the addition of alpha 1 to the complex. This finding, together with the observation that Q-element point mutations affected ternary complex formation but not the weak binding of MCM1 alone, supports the idea that the Q sequence serves as a binding site for alpha 1.

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