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At least 397 records · Page 22Linked to original sources
PubMed1982

Selective binding of actinomycin D and distamycin A to DNA.

The exact sites at which a number of drugs inhibit the nick translation of DNA by E.coli DNA polymerase-I have been pinpointed. In order to do this, a method has been developed for sequencing double-stranded plasmid DNA from the site of a specifically induced nick. The initial experiments have concentrated on analysis of drug inhibition of nick translation in a 200 nucleotide region near the Eco Rl origin of pBR313. Many drugs were found to inhibit nick translation in a highly sequence specific manner. For actinomycin D, significant inhibition occurred at just four sites in the nucleotide sequence under test and only one sequence (pGpCpGpCpGpGp) gave really strong inhibition. Distamycin A gave a different pattern of inhibition with particularly strong stops in just two of the many A-T rich regions in the DNA. Experiments with caffeine suggest that factors in addition to primary sequence are important in determining where major inhibition occurs.

Adenoviruses, Human↗
PubMed1999

Detection of mitochondrial DNA mutations by temporal temperature gradient gel electrophoresis.

BACKGROUND: A unique requirement for the molecular diagnosis of mitochondrial DNA (mtDNA) disorders is the ability to detect heteroplasmic mtDNA mutations and to distinguish them from homoplasmic sequence variations before further testing (e.g., sequencing) is performed. We evaluated the potential utility of temporal temperature gradient gel electrophoresis (TTGE) for these purposes in patients with suspected mtDNA mutations. METHODS: DNA samples were selected from patients with known mtDNA mutations and patients suspected of mtDNA disorders without detectable mutations by routine analysis. Six regions of mtDNA were PCR amplified and analyzed by TTGE. Electrophoresis was carried out at 145 V with a constant temperature increment of 1.2 degrees C/h. Mutations were identified by direct sequencing of the PCR products and confirmed by PCR/allele-specific oligonucleotide or PCR/restriction fragment length polymorphism analysis. RESULTS: In the experiments using patient samples containing various amounts of mutant mtDNA, TTGE detected as little as 4% mutant heteroplasmy and identified heteroplasmy in the presence of a homoplasmic polymorphism. In 109 specimens with 15 different known mutations, TTGE detected the presence of all mutations and distinguished heteroplasmic mutations from homoplasmic polymorphisms. When 11% of the mtDNA genome was analyzed by TTGE in 104 patients with clinically suspected mitochondrial disorders, 7 cases of heteroplasmy ( approximately 7%) were detected. CONCLUSIONS: TTGE distinguishes heteroplasmic mutation from homoplasmic polymorphisms and appears to be a sensitive tool for detection of sequence variations and heteroplasmy in patients suspected of having mtDNA disorders.

Alleles↗
PubMed1996

Ankylosing spondylitis and heterogeneity of HLA-B27 in Chinese.

OBJECTIVE: To examine the relationship between ankylosing spondylitis and heterogeneity of HLA-B27 in Chinese. PATIENTS AND METHODS: We subtyped HLA-B27 in 76 patients with ankylosing spondylitis and 1 200 healthy Chinese of Han nationality. Polymerase chain reaction was designed to amplify specific regions of class I (exon 2 and 3). Amplified sequences were tested with eight sequence-specific oligonucleotides to distinguish all HLA-B27 subtypes. RESULTS: The HLA-B27 was closely related to ankylosing spondylitis and at least four subtypes presented in Chinese patients. B2704 was the most common subtype, occurring in 54.8% and 50.85% respectively in HLA-B27 positive ankylosing spondylitis patients and healthy controls. The main HLA-B27 subtypes in the HLA-B27 positive patients distributed in similar proportion to those of HLA-B27 positive healthy controls (P < 0.1). CONCLUSIONS: The prevalences of various subtypes of HLA-B27 vary in different races. HLA-B2704 is the main subtype in Chinese Han nationality. The HLA-B27 specific residues that are shared by the subtypes might be involved in the pathogenesis of ankylosing spondylitis.

Adolescent↗
PubMed1996

[Ankylosing spondylitis and heterogenity of HLA-B27 in Chinese].

HLA-B27 represents a family of closely related antigens. More than six alleles differing in limited number of nucleotide substitution had been described (B2701-B2706). We subtyped HLA-B27 in 76 patients with ankylosing spondylitis (AS) and 1200 Chinese healthy persons of Han nationality. Polymerase chain reaction was designed to amplify specific regions for class I (exon 2 and 3). Amplified sequences were tested with eight sequence-specific oligonucleotides to distinguish all HLA-B27 subtypes. The results showed: the HLA-B27 was closely related with AS and at least four subtypes existed in Chinese patients. B2704 was the most common subtype occurring in 54.8% and 50.85% respectively in HLA-B27 positive AS patients and healthy controls. The main HLA-B27 subtypes in the HLA-B27 positive patients distributed in similar proportion to that of HLA-B27 positive healthy controls (P > 0.1). We believe that the above results imply that the prevalences of the various subtypes of HLA-B27 vary in different races. The HLA-B27 specific residues that are shared by the subtypes might be involved in the pathogenesis of AS.

Alleles↗
PubMed1995

Use of maximum length sequence analysis in newborn hearing testing.

The use of maximum length sequence analysis (MLSA) with rapid click rates may be of clinical value in newborn hearing screening because a greater number of individual responses can be signal averaged without adding to test time. To examine the potential clinical value of MLSA in newborn screening, auditory brainstem responses (ABRs) from 50 premature newborns were studied. ABRs were acquired with conventional signal averaging at four stimulus intensity levels (50 dB, 40 dB, 30 dB, and 20 dB nHL) using a click rate of 33.3/sec. These responses were directly compared with the ABRs acquired with MLSA using a rate of 227.3/sec. MLSA and conventional signal averaging yielded similar results with no statistically significant differences in the number of responses detected. Across babies, the overall quality of the tracings slightly favored MLSA, particularly when recording conditions were poor. Though the results of this investigation do not support the use of MLSA as the primary technique in newborn screening in the neonatal intensive care unit (NICU), they do support consideration of the use of MLSA as an alternative technique when the responses obtained with conventional signal averaging are poorly defined.

Acoustic Stimulation↗
PubMed1993

Single-strand conformational polymorphism and direct sequencing applied to carrier testing in families with ornithine transcarbamylase deficiency.

Single-strand conformational polymorphism (SSCP) and direct sequencing were used to confirm or deny carrier status in three families with ornithine transcarbamylase (OTC) enzyme deficiency. Two male probands with "late onset" OTC deficiency, whose "private" mutations were previously characterized, inherited the mutations form their heterozygous mothers. One of the heterozygous mothers had a false negative allopurinol test. Three female siblings of the two male probands were tested, one proved to be a carrier of the respective mutation while the other two were found to have normal alleles. In the third family, the proband was a female with "late onset" presentation of OTC deficiency. We found a new point mutation in this girl consisting of a guanine-to-cytosine transversion at nucleotide 520 resulting in a substitution of proline for alanine at amino acid 142 of the mature OTC protein. We confirmed that this mutation occurred spontaneously and that neither of the two parents carries this mutation. We conclude that SSCP, in conjunction with direct sequencing, is a useful technique that can be practically applied for carrier testing in families with OTC deficiency.

Amino Acid Metabolism, Inborn Errors↗