Isothermal Amplification Methods for the SNP Genotyping

2019 ◽  
Vol 19 (7) ◽  
pp. 461-472 ◽  
Author(s):  
Somayeh Heidari Sharafdarkolaee ◽  
Pooria Gill ◽  
Majid Motovali-Bashi ◽  
Fatemeh Heidari Sharafdarkolaee
Keyword(s):  
Mutation Detection ◽  
Driving Forces ◽  
Snp Genotyping ◽  
Isothermal Amplification ◽  
Chain Reaction ◽  
Novel Technologies ◽  
Detection Techniques ◽  
Wide Range ◽  
Polymerase Chain ◽  
Thermal Cycler

The demands for genotyping techniques with acceptable precision, accuracy, cost-effectiveness in high throughput formats made driving forces for continuous development of novel technologies. A wide range of mutation detection techniques based on polymerase chain reaction (PCR) have been introduced. The best alternatives were the isothermal amplification technologies that those did not require a thermal cycler. In this review, we aimed to describe the most known isothermal amplification techniques for SNP genotyping.

A Prototype Continuous Flow Polymerase Chain Reaction LTCC Device

2007 ◽  
Vol 539-543 ◽  
pp. 523-528 ◽  
Author(s):  
Korey Moeller ◽  
Jason Besecker ◽  
Greg Hampikian ◽  
A. Moll ◽  
D. Plumlee ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Low Temperature ◽  
Continuous Flow ◽  
Low Cost ◽  
Flow Characteristics ◽  
Chain Reaction ◽  
Wide Range ◽  
Polymerase Chain ◽  
Thermal Cycler ◽  
Biological Entities

There is a growing need for remote biological sensing in both laboratory and harsh field environments. Sensing and detection of biological entities such as anthrax, Ebola and other micro-organisms of interest involves sampling of the environment, amplification, analysis and identification of the target DNA. A key component of such a sensor is a low cost, portable, reusable, continuous flow polymerase chain reaction (PCR) thermal cycler. Fabrication with low temperature co-fired ceramics (LTCC) can provide a reusable low cost device capable of operating in a wide range of environments The design and manufacture of a prototype continuous flow micro-fluidic PCR device using low temperature co-fired ceramic is presented. Initial modeling of flow characteristics and heat transfer was carried out in SolidWorks™. The prototype device employs resistance heaters below the channels, buried and surface thermocouples for temperature monitoring, and air gaps for thermal isolation.

scholarly journals Loop-Mediated Isothermal Amplification (LAMP): Comparative Advances over Conventional PCR and Other Molecular Techniques

2020 ◽  
pp. 33-44
Author(s):  
Yahaya Hassan ◽  
Leslie Thian Lung Than
Keyword(s):  
Target Genes ◽  
Molecular Techniques ◽  
Isothermal Amplification ◽  
Nucleic Acid Amplification ◽  
Conventional Pcr ◽  
Chain Reaction ◽  
Loop Mediated Isothermal Amplification ◽  
Naked Eye ◽  
Polymerase Chain ◽  
Thermal Cycler

Gene amplification technology is essential in the fields of diagnostic medicine. The polymerase chain reaction (PCR) is central in the molecular studies and provides ways for diagnostic advancement in the areas. However, the requirement for thermal cycler in a dedicated facility for amplification of target genes in the PCR technique has been a bottleneck to many researchers. The limitations associated with PCR include cost implication, strict expertise necessity and relatively higher turn-around time. The emergence of loop-mediated isothermal amplification (LAMP) in the last two decades assists in bridging the undesirable gaps. This review aims to highlight the natural advantages of the LAMP technique over the existing conventional PCR and other isothermal molecular techniques. Available published articles on LAMP techniques reviewed, listed many outstanding advances of the method in comparison to traditional PCR technique. The mentioned advantages include simplicity, affordability, naked-eye result detection and many more. That made LAMP becomes a rapidly accepted method in the field of molecular diagnosis. Other essential features of LAMP in comparison with other emerging nucleic acid amplification techniques were adequately explained and presented in tabular form for research and quick reference purposes. Though LAMP has some few limitations, its advantages outweigh its flaws by filling the gap in the field of molecular biology diagnostics.

scholarly journals Comparative Study of Immune-Diagnostic Tools with Polymerase Chain Reaction in Sub-Clinical Leishmaniasis Isolates

2011 ◽  
Vol 12 (1) ◽  
pp. 34-39 ◽  
Author(s):  
Nazar M Abdalla
Keyword(s):  
Polymerase Chain Reaction ◽  
Skin Test ◽  
Diagnostic Methods ◽  
Diagnostic Tools ◽  
Enzyme Linked Immunosorbent Assay ◽  
Clinical Form ◽  
Chain Reaction ◽  
Leishmanin Skin Test ◽  
Wide Range ◽  
Polymerase Chain

Objective: This study aimed to identify cases of leishmaniasis in the Nuba Mountain area, which is situated in a unique geographical site located in the centre of Sudanese leishmania belt. Wide range of investigations are available for detection of leishmania cases, but still the most reliable and easy test used as screening and epidemiological tool in field studies needs to be evaluated. The most commonly used conventional diagnostic methods direct microscopy and culture have some drawbacks in diagnosing subclinical cases of leishmaniasis. Materials and methods: In this study, comparative properties of various immune-diagnostic tools with Polymerase Chain Reaction used in sub-clinical leishmaniasis isolates were explored. The immune-diagnostic tools involved in this study include- Leishmanin Skin Test (LST), Enzyme Linked Immunosorbent Assay (ELISA) and Direct Agglutination Test (DAT). The study was conducted in the Green Valley village (Rashad Province, South Kordofan State) with a population of 332. Most of the villagers presented with sub-clinical form of leishmaniasis with minor symptoms and signs without the features found in clinical form of visceral leishmaniasis such as fever, diarrhoea, epistaxis, enlarged lymph nodes, spleen and liver. In this study we collected demographic, clinical and epidemiological data using special questionnaire. Leishmanin skin test (LST), ELISA, DAT and PCR for parasite DNA detection were used. Result: The final positive cases detected by PCR were 32 out of 332 belong to L. donovani species. The final positive cases detected by LST were 51.2% of the total population under study, while 11 out of the 37 tested samples were positive by ELISA. All of the 332 villagers showed negative readings by DAT with exception of three individuals who were positive with very high titers. Conclusion: DNA etxtraction and amplification with primers can be a good screening tool in subclinical leishmaniasis isolates. Keyword: Sub-clinical; Leishmaniasis; Leishmanin Skin Test; ELISA; DAT; PCR. DOI: 10.3329/jom.v12i1.5422J Medicine 2011; 12 : 34-39

scholarly journals Design and development of polymerase chain reaction thermal cycler using proportional-integral temperature controller

2018 ◽  
Vol 14 (2) ◽  
pp. 213-218
Author(s):  
Chong Kim Soon ◽  
Nawoor Anusha Devi ◽  
Kok Beng Gan ◽  
Sue-Mian Then
Keyword(s):  
Polymerase Chain Reaction ◽  
Low Cost ◽  
Maximum Temperature ◽  
Gradient System ◽  
Chain Reaction ◽  
Temperature Controller ◽  
Proportional Integral ◽  
Integral Temperature ◽  
Polymerase Chain ◽  
Thermal Cycler

A thermal cycler is used to amplify segments of DNA using the polymerase chain reaction (PCR). It is an instrument that requires precise temperature control and rapid temperature changes for certain experimental protocols. However, the commercial thermal cyclers are still bulky, expensive and limited for laboratory use only.  As such it is difficult for on-site molecular screening and diagnostics. In this work, a portable and low cost thermal cycler was designed and developed. The thermal cycler block was designed to fit six microcentrifuge tubes. A Proportional-Integral temperature controller was used to control the thermal cycler block temperature. The results showed that the maximum temperature ramp rate of the developed thermal cycler was 5.5 °C/s. The proportional gain (Kp) and integral gain (Ki) of the PI controller were 15 A/V and 1.8 A/Vs respectively. Finally, the developed thermal cycler successfully amplified six DNA samples at the expected molecular weight of 150 base pair. It has been validated using the Eppendorf Mastercycler nexus gradient system and gel electrophoresis analysis

scholarly journals Comparison of a PCR-based diagnostic assay for Mycoplasma pulmonis with traditional detection techniques

1994 ◽  
Vol 28 (3) ◽  
pp. 249-256 ◽  
Author(s):  
S. Sanchez ◽  
K. Tyler ◽  
N. Rozengurt ◽  
J. Lida
Keyword(s):  
Diagnostic Methods ◽  
Diagnostic Assay ◽  
Mycoplasma Pulmonis ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Culture Techniques ◽  
Detection Techniques ◽  
Laboratory Rodent ◽  
Polymerase Chain ◽  
Diagnosis Of Infection

Current diagnosis of infection by Mycoplasma pulmonis, an important pathogen of laboratory rodent colonies worlwide, is based on serological, histopathological and culture techniques which can be slow and unreliable. A polymerase chain reaction (PCR) assay for M. pulmonis diagnosis was compared to current diagnostic methods. This PCR based technique allows a more specific, sensitive and rapid diagnosis of M. pulmonis from various tissues by comparison with culture and histopathology.

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