Restriction fragment length polymorphisms of the apolipoprotein A-I, C-III, A-IV gene locus Relationships with lipids, apolipoproteins, and premature coronary artery disease
José M. OrdovásFernando CiveiraJacques GenestS. E. CraigA H RobbinsT MEADEMiguel Pocovı́Philippe M. FrossardU. MasharanPeter W.F. WilsonDeeb N. SalemR. H. WardErnst J. Schaefer
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Abstract:
Data from various laboratories have indicated associations of various alleles determined by RFLPs within or adjacent to several apolipoprotein genes with abnormalities in plasma lipids and/or premature coronary artery disease (CAD). In order to assess such relationships we have examined allele frequencies of 8 different RFLPs within or adjacent to the apo A-I, C-III and A-IV gene complex on the long arm of chromosome 11 (MspI, 5′ to the apo A-I gene; MspI, within the apo A-1 gene; PstI, 3′ to the apo A-I gene; SstI, 3′ to the apo C-III gene; PvuII, within the apo C-III gene; PvuII, 5′ to the apo C-III gene; XbaI, within the apo A-IV gene; and XbaI, 3′ to the apo A-IV gene) in 202 patients with CAD (50% narrowing of one or more coronary arteries) prior to age 60 and 145 normal controls. None of the allele frequencies of these RFLPs were significantly different in cases as compared to controls. With regard to associations with plasma lipids and apolipoprotein levels, the rare allele determined by the absence of the PstI site was associated with elevated triglyceride levels (P < 0.05) in cases, but not in controls. In contrast, the rare MspI allele 5′ to the apo A-I gene was associated with elevated triglyceride levels (P < 0.05) in controls but not in cases. In both cases and controls, subjects with the uncommon SstI allele had triglyceride levels that were 9 and 38% higher than in those without this allele. These differences were significant (P < 0.05) only in controls. Our data indicate that the rare allele determined by the SstI site within this gene complex deserves further study in order to understand its association with elevated triglycerides in Caucasian populations. However, at the present time all these DNA markers lack sufficient specificity to be clinically useful for CAD risk assessment.Keywords:
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Apart from birds, Mycobacterium avium subsp. avium (MAA) is often isolated from granulomatous lesions in pigs and occasionally from cattle and other animals. The objectives of this study were the detection of IS901 restriction fragment length polymorphism (RFLP) types of MAA isolates from different species of domestic animals between the years 1998 and 2004 and the comparison of the detected RFLP types with previously described RFLP types collected in the database of the OIE Reference Laboratory for Avian Tuberculosis (Brno, Czech Republic). Furthermore, the RFLP types of the isolates obtained from MAA outbreaks on one of the largest pig farms in Slovenia were also investigated. A total of 62 isolates (56 from pigs, five from poultry and one from cattle) were identified with IS901 PCR and IS901 RFLP typed using restriction endonucleases PvuII and PstI. Seven PvuII RFLP and 11 PstI RFLP types resulted in 12 combined PvuII PstI types; none of these matched the combined RFLP types described in previous studies. Our contributions to the database were two new PvuII and eight new PstI RFLP types. Identical RFLP types were found among isolates of animals originating from individual farms. Finding of identical RFLP types within a farm is not surprising because the animals were epidemiologically related and infected with one strain. A unique RFLP type F-A17 was detected in isolates from different pig herds and also in isolates from poultry. Detection of identical RFLP types on different farms may reflect one MAA source. The other combined PvuII PstI RFLP types were identified only once which indicates considerable variety of MAA RFLP types in Slovenia.
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Research about PCR-RFLP gene 16S rRNA in toad (Bufonidae) using restriction ezymes has conducted from October 2016 to January 2017. The aim of this research is to detect variation from three species of Bufonidae using PCR-RFLP method gene 16S rRNA. DNA sampel was isolated dan amplified by PCR process. RFLP was done using two restriction enzymes, MseI (Tru1I) and HphI. For restriction enzyme MseI produce different restriction sites and fragment that are vary in length for three species of toad that makes us able to detect variation. RFLP with restriction enzymes HphI produce restriction sites and fragments that are equal in length so it can’t be use to detect variation between three species of toad from genus Duttaphrynus and Phrynoidis
Keywords: Bufonidae, PCR-RFLP, 16S rRNA, primer, restriction enzyme
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Herpesvirus salmonis (H. salmonis) and seven strains of Oncorhynchus masou virus (OMV) were analyzed for their genomes with restriction endonucleases HindIII and PstI. Restriction profiles of H. salmonis were different from those of OMV strains. Restriction patterns of OMV strain DNAs were divided into four subtypes with HindIII. With PstI digestion, DNA restriction patterns of those strains were divided into five subtypes. Obvious difference in restriction patterns was not observed among tumor tissue-derived and coelomic fluids-derived strains. By using 32P-labelled DNA of standard OMV (strain 00-7812) as a probe, H. salmonis DNA was not hybridized, but most of the fragments of OMV strain DNAs were hybridized.
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Abstract Gupta, P.K. and Rai, A. 1994. Restriction map of cloned 3.8 kb PstI fragment of bovine herpes virus-1 DNA. J. Appl. Anim. Res., 6: 143–149. A 3.8 kb Pstl fragment of bovine herpesvirus-1 DNA was selectively cloned into pUC9 plasmid vector. The recombinant plasmid with the insert was analyzed using restriction endonucleases. On the basis of restriction digested fragment pattern, partial restriction site map was prepared for a few restriction endonucleases. Shot gun cloning of PstI digested DNA fragments yielded two clones having double inserts.
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Thirty four taxa of Actinidia were investigated through PCR RFLP analysis of two mitochondrial DNA fragments, including nad1/B C gene and nad4/1 2 gene. Digestion of these two fragments by seven restriction endonucleases yielded twenty polymorphic restriction fragments. We only found restriction fragment length variation caused by insertion or deletion. It suggests that mtDNA restriction length polymorphism could be an efficient indicator for conservation biology .
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The genomic variation of three isolates of bovine herpes virus 1 (BHV-1) originating from different geographical regions and displaying different clinical symptoms were studied by restriction analysis using four different restriction endonucleases. EcoRI displayed an uniform restriction pattern for all the isolates and HindIII showed marginal variation among the isolates. BstEII and PstI displayed unique restriction patterns, based on which the isolates could be grouped into two subgroups of BHV-1.1. BstEII appears to be the enzyme of choice for differentiating BHV-1.1.
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SUMMARY The DNA of a field isolate and of a vaccine strain of canine adenovirus type 2 (CAV-2) were analysed by digestion with several restriction endonucleases. The PstI restriction fragments of the field isolate (CAV-2 Glasgow) and the vaccine strain were cloned into the plasmid pBR322. Physical maps of the two viral genomes were constructed by molecular hybridization of PstI, EcoRI, SmaI, BamHI and KpnI digests of the viral DNA with the cloned PstI fragments. The restriction profile of CAV-2 Glasgow was shown to be virtually identical to those of the two prototype CAV-2 strains, Toronto A26/61 and Manhattan. However, the restriction fragment pattern of the vaccine strain of CAV-2 showed characteristic alterations, in particular additional sequences at or near the genome termini.
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Summary IS1245-based restriction fragment length polymorphism (RFLP) analysis has been proposed recently for molecular typing of Mycobacterium avium isolates. As there is no standardised method with respect to the optimal restriction enzyme, three restriction endonucleases were tested for analysis of 17 human isolates. The restriction endonucleases, selected on the basis of the physical maps of IS1245 and of the highly homologous IS1311, were BsaAI, that cleaves IS1245, PvuII, that cleaves IS1311, and NruI, that cleaves both IS1245 and IS1311. All the restriction endonucleases yielded polymorphic and complex RFLP patterns. However, BsaAI- and NruI-generated bands were more evenly distributed and easier to detect than PvuII-generated bands, most of which clustered in a narrow zone of the fingerprint. In some cases, DNA digestion with BsaAI or NruI yielded probe-specific restriction fragments of molecular size lower than expected. Moreover, digestion with NruI, which was expected to generate the highest numbers of bands in all the isolates, yielded fewer bands than were obtained with BsaAI or PvuII in 14 and 5 isolates, respectively. These findings might suggest the existence of unidentified IS1245-related insertion element(s) in M. avium isolates. Computer analysis of the IS1245-based RFLP patterns of M. avium isolates showed that the restriction endonucleases were capable, although with minor differences, of defining distinct banding patterns and clusters of identical or highly related isolates, thus confirming IS1245-based RFLP analysis as a useful technique for epidemiological studies.
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Polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analysis were used to differentiate between serotypes of several infectious bronchitis virus (IBV) strains. A sequence of 1720 base pairs (bp) that contains the S1 glycoprotein gene of IBV was amplified by PCR, purified, and digested with restriction enzymes. Eleven reference IBV strains were grouped according to the RFLP patterns. The IBV Holte, Arkansas DPI, SE 17, Md 27, and Iowa 97 strains could be differentiated from the other IBV strains using the restriction enzyme HaeIII. The Beaudette, Massachusetts 41, Connecticut, and Florida 88 strains had the same HaeIII RFLP pattern but could be differentiated using XcmI and BstYI restriction enzymes. The Gray and JMK strains could not be differentiated by their RFLP patterns following digestion with 23 different restriction enzymes. Twenty-six samples (field isolates and reference strains) of IBV, previously serotypes by the virus-neutralization (VN) test in embryonating eggs, were analyzed in a blind fashion. The results using the PCR and RFLP analysis agreed with the serotype for traditional and variant IBV viruses as determined by the VN test.
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