scholarly journals LARGE-SCALE, POLYMERASE CHAIN REACTION–BASED SURVEILLANCE OF SCHISTOSOMA HAEMATOBIUM DNA IN SNAILS FROM TRANSMISSION SITES IN COASTAL KENYA: A NEW TOOL FOR STUDYING THE DYNAMICS OF SNAIL INFECTION

2004 ◽  
Vol 71 (6) ◽  
pp. 765-773 ◽  
Author(s):  
JOSEPH HAMBURGER ◽  
ORIT HOFFMAN ◽  
H. CURTIS KARIUKI ◽  
ERIC M. MUCHIRI ◽  
CHARLES H. KING ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Schistosoma Haematobium ◽  
Large Scale ◽  
Snail Infection ◽  
Chain Reaction ◽  
Coastal Kenya ◽  
Polymerase Chain

Pitfalls in the epidemiologic classification of human papillomavirus types associated with cervical cancer using polymerase chain reaction: driver and passenger

2008 ◽  
Vol 18 (5) ◽  
pp. 1042-1050 ◽  
Author(s):  
T. Matsukura ◽  
M. Sugase
Keyword(s):  
Cervical Cancer ◽  
Polymerase Chain Reaction ◽  
Human Papillomavirus ◽  
Large Scale ◽  
Blot Hybridization ◽  
Southern Blot Hybridization ◽  
Cervical Lesions ◽  
Chain Reaction ◽  
Polymerase Chain

Cervical cancer is a common malignancy in women worldwide, and it has now been established that the human papillomavirus (HPV) is both necessary and causal for these lesions. HPV itself is both ubiquitous and markedly heterogeneous but can nevertheless be classified as either a high-risk type or a low-risk type based upon its frequency of detection in cervical cancer. Given that the association between HPV and cervical cancer is causal, the classification of this virus has been strengthened by large-scale epidemiologic studies and is widely accepted across many disciplines. It is evident, however, that cervical cancer is frequently associated with multiple HPV types. Therefore, it is crucial to distinguish causal types of HPV (drivers) from noncausal types (passengers) in cervical lesions. In this review, we highlight the current pitfalls of using polymerase chain reaction methods instead of Southern blot hybridization for detecting HPV and discuss the distinction between driver and passenger HPVs with regard to the viral type, the length of the viral genome, and the levels of viral DNA associated with cervical cancer. Finally, we newly propose three categories of HPV instead of two risk groups, based on similarities between viral genes

Metagenomics to Paleogenomics: Large-Scale Sequencing of Mammoth DNA

Science ◽  
2005 ◽  
Vol 311 (5759) ◽  
pp. 392-394 ◽  
Author(s):  
Hendrik N. Poinar ◽  
Carsten Schwarz ◽  
Ji Qi ◽  
Beth Shapiro ◽  
Ross D. E. MacPhee ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Dna Sequence ◽  
Large Scale ◽  
Loxodonta Africana ◽  
Chain Reaction ◽  
Base Pairs ◽  
Mammuthus Primigenius ◽  
Sequence Identity ◽  
Polymerase Chain ◽  
Woolly Mammoth

We sequenced 28 million base pairs of DNA in a metagenomics approach, using a woolly mammoth (Mammuthus primigenius) sample from Siberia. As a result of exceptional sample preservation and the use of a recently developed emulsion polymerase chain reaction and pyrosequencing technique, 13 million base pairs (45.4%) of the sequencing reads were identified as mammoth DNA. Sequence identity between our data and African elephant (Loxodonta africana) was 98.55%, consistent with a paleontologically based divergence date of 5 to 6 million years. The sample includes a surprisingly small diversity of environmental DNAs. The high percentage of endogenous DNA recoverable from this single mammoth would allow for completion of its genome, unleashing the field of paleogenomics.

Detection ofTrypanosoma congolenseandTrypanosoma bruceisubspecies by DNA amplification using the polymerase chain reaction

Parasitology ◽  
1989 ◽  
Vol 99 (1) ◽  
pp. 57-66 ◽  
Author(s):  
D. R. Moser ◽  
G. A. Cook ◽  
Diane E. Ochs ◽  
Cheryl P. Bailey ◽  
Melissa R. McKane ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Dna Sequences ◽  
Large Scale ◽  
Nuclear Dna ◽  
Pcr Amplification ◽  
Dna Amplification ◽  
Detection Methods ◽  
Chain Reaction ◽  
African Trypanosomes ◽  
Polymerase Chain

SUMMARYThe nuclear DNA ofTrypanosoma congolensecontains a family of highly conserved 369 base pair (bp) repeats. The sequences of three cloned copies of these repeats were determined. An unrelated family of 177 bp repeats has previously been shown to occur in the nuclear DNA ofTrypanosoma brucei brucei(Sloofet al.1983a). Oligonucleotides were synthesized which prime the specific amplification of each of these repetitive DNAs by the polymerase chain reaction (PCR). Amplification of 10% of the DNA in a single parasite ofT. congolenseorT. bruceispp. produced sufficient amplified product to be visible as a band in an agarose gel stained with ethidium bromide. This level of detection, which does not depend on the use of radioactivity, is about 100 times more sensitive than previous detection methods based on radioactive DNA probes. The oligonucleotides did not prime the amplification of DNA sequences in other trypanosome species nor inLeishmania, mouse or human DNAs. Amplification of DNA from the blood of animals infected withT. congolenseand/orT. bruceispp. permitted the identification of parasite levels far below that detectable by microscopic inspection. Since PCR amplification can be conducted on a large number of samples simultaneously, it is ideally suited for large-scale studies on the prevalence of African trypanosomes in both mammalian blood and insect vectors.

A polymerase chain reaction assay for cucumber mosaic virus in lupin seeds

1993 ◽  
Vol 44 (1) ◽  
pp. 41 ◽  
Author(s):  
S Wylie ◽  
CR Wilson ◽  
RAC Jones ◽  
MGK Jones
Keyword(s):  
Polymerase Chain Reaction ◽  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Large Scale ◽  
Seed Infection ◽  
Chain Reaction ◽  
Routine Testing ◽  
Source Of Infection ◽  
Lupin Seed ◽  
Polymerase Chain

Seed is the main source of infection of narrow-leafed lupin (Lupinus angustifolius) crops by cucumber mosaic virus (CMV). The ELISA procedure is currently used for large-scale, routine testing of lupin seed samples, but a more sensitive, reliable and labour-saving assay is needed which detects levels of seed infection as low as 0.1%. A Polymerase Chain Reaction (PCR) using ground dry seed samples was developed for this purpose. Primers based on concensus sequences of eight published CMV coat protein cDNAs (RNA3) of CMV subgroups 1 and 2 were used. The assay involved (1) a reverse transcription step for cDNA synthesis and (2) amplification of a specific fragment (482-501 bp depending on the strain) by PCR. Two methods of extracting virus from infected lupin material were used: (i) a rapid procedure which was effective for samples with higher levels of infection, e.g. infected leaves and 20.5% infected seed; (ii) a phenol-chloroform procedure, which led to greater sensitivity, enabling reliable detection of 0.1% seed infection. It detected CMV in 16 commercial seed samples (0.1-8% seed infection) belonging to seven cultivars from 12 different localities. Both methods were suitable for routine testing of the flour derived from grinding dry seed. On dissection of infected seeds, CMV was detected in the cotyledons and embryo and usually in or on the testa. The PCR assay detected virus from both CMV subgroups, but only subgroup 2 was found in lupin seed samples. The two CMV subgroups can be distinguished by digestion of amplified DNA with the restriction enzyme EcoRI; only CMV strains of subgroup 2 are digested to yield two fragments of size 330 and 170 bp.

Nanotechnology in medicine. The use of polymerase chain reaction in the diagnosis of infectious diseases

2020 ◽  
pp. 23-29
Author(s):  
Vladimir Anatolievich Klimov
Keyword(s):  
Polymerase Chain Reaction ◽  
Infectious Diseases ◽  
Large Scale ◽  
Medical Science ◽  
Laboratory Diagnostics ◽  
Protein Markers ◽  
Chain Reaction ◽  
Dna Molecule ◽  
Stage Of Development ◽  
Polymerase Chain

At all times mankind strived for progressive changes, and for a long time has been looking for new ways to diagnose and treat diseases in order to prolong life. Scientists have constantly attempted to introduce the latest techniques, and if in the 20th century an understanding of the course of various processes at the cellular level was achieved, at the latest stage of development there is already a transition to the study of the molecular and even atomic composition of individual derivatives, which should contribute to the transition to a qualitatively new level of process understanding. The first to speak about nanotechnology was Richard Phillips Feynman, who back in 1959 spoke about the possibility of controlling matter precisely at the atomic level. At the present stage, nanotechnology is increasingly being introduced into medical science, in particular, in the field of laboratory diagnostics of infectious diseases. More recently, this fact has received practical confirmation on the example of organizing large-scale testing of the population for the presence of the coronavirus infection. The methods used on the basis of atomic force molecular detectors provide a unique opportunity for visualization and identification of protein markers of pathological processes and conditions with a sensitivity several orders of magnitude higher than that of standard laboratory studies. This principle was the basis for the implementation of the polymerase chain reaction method, the essence of which lies in the multiple multiplication of microscopic concentrations of pathogen DNA fragments in a patient’s biological sample under artificial conditions. As a result of a complex process called amplification, under the influence of enzymes and changes in temperature (from 50 to 95 °C), two DNA molecules are formed from one DNA molecule. In this case, there is a copying of a DNA section that is present only in that type of pathogenic microorganism that is of interest to a specialist at the moment. The cycle of formation of a new DNA molecule takes about 3 minutes, while 30-40 cycles is quite enough to obtain the proper number of molecules required for reliable visual determination of the desired agent by electrophoresis.

scholarly journals Prevalence of human T-lymphotropic virus 1/2 in Nigeria’s capital territory and meta-analysis of Nigerian studies

2019 ◽  
Vol 7 ◽  
pp. 205031211984370
Author(s):  
Nneoma Confidence JeanStephanie Anyanwu ◽  
Elijah Ekah Ella ◽  
Maryam Aminu ◽  
Maleeha Azam ◽  
Muhammad Ajmal ◽  
...  
Keyword(s):  
Risk Factors ◽  
Polymerase Chain Reaction ◽  
Immunosorbent Assay ◽  
Large Scale ◽  
Meta Analysis ◽  
Antibody Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Chain Reaction ◽  
T Lymphotropic Virus ◽  
Polymerase Chain

Objectives: This study was aimed at determining human T-lymphotropic virus 1/2 prevalence among apparently healthy, immunocompromised and haematologic malignant individuals in Nigeria’s capital, as well as meta-analysis of all Nigerian studies until date. Methods: A total of 200 participants were recruited into a cross-sectional study. In total, 1 mL each of sera and plasma were obtained from 5 mL blood of each participant and analysed for antibodies to human T-lymphotropic virus 1/2 using enzyme-linked immunosorbent assay; positive samples confirmed with qualitative real-time polymerase chain reaction, followed by statistical and meta-analysis. Sociodemographic characteristics and possible risk factors were assessed via questionnaires. Results: Enzyme-linked immunosorbent assay yielded 1% prevalence which was confirmed to be zero via polymerase chain reaction. A total of 119 (59.5%) of the participants were male, while the mean age was 35.28 ± 13.61 years. Apart from sex and blood reception/donation, there was generally a low rate of exposure to human T-lymphotropic virus–associated risk factors. Meta-analysis revealed pooled prevalence of human T-lymphotropic virus 1 and 2 to be 3% and 0%, respectively, from Nigerian studies. Conclusion: This study discovered zero prevalence of human T-lymphotropic virus 1/2 from five major hospitals in Nigeria’s capital, exposing the importance of confirmatory assays after positive antibody detection assay results. Meta-analysis highlighted the existence of very few reliable Nigerian studies compared to the demography of the nation. Large-scale epidemiological studies and routine screening of risk populations are therefore needed since Nigeria lies in the region of endemicity.

AUTOMATED VALIDATION OF POLYMERASE CHAIN REACTION AMPLICON MELTING CURVES

2006 ◽  
Vol 04 (02) ◽  
pp. 299-315
Author(s):  
TOBIAS P. MANN ◽  
RICHARD HUMBERT ◽  
JOHN A. STAMATOYANNOPOLOUS ◽  
WILLIAM STAFFORD NOBLE
Keyword(s):  
Polymerase Chain Reaction ◽  
Large Scale ◽  
Fundamental Problem ◽  
Support Vector ◽  
Chain Reaction ◽  
Single Product ◽  
Widespread Application ◽  
Major Barrier ◽  
Melting Curves ◽  
Polymerase Chain

The polymerase chain reaction (PCR) is a fundamental tool of molecular biology. Quantitative PCR is the gold-standard methodology for determination of DNA copy numbers, quantitating transcription, and numerous other applications. A major barrier to large-scale application of PCR for quantitative genomic analyses is the current requirement for manual validation of individual PCRs to ensure generation of a single product. This typically requires visual inspection either of gel electrophoreses or temperature dissociation ("melting") curves of individual PCRs — a time-consuming and costly process. Here we describe a robust computational solution to this fundamental problem. Using a training set of 10 080 reactions comprising multiple quantitative PCRs from each of 1728 unique human genomic amplicons, we developed a support vector machine classifier capable of discriminating single-product PCRs with better than 99% accuracy. This approach has broad utility, and eliminates a major bottleneck to widespread application of PCR for high-throughput genomic applications.

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