scholarly journals Comparison of Three Strategies for Sequencing Rabies Viral G Gene

2013 ◽  
Vol 2 (1) ◽  
Author(s):  
Shengli Meng ◽  
Wenli Lv ◽  
Jie Wu ◽  
Jilin Wang ◽  
Gelin Xu ◽  
...  
Keyword(s):  
Direct Sequencing ◽  
Low Cost ◽  
Cost Effective ◽  
Brain Homogenate ◽  
Universal Primers ◽  
Viral Gene ◽  
Gene Sequences ◽  
Rt Pcr ◽  
Pcr Method ◽  
Polymerase Chain

<p>It is essential to rapidly, low-cost and precisely determining gene sequences of rabies virus. Reverse transcription-polymerase chain reaction can be used to identify rabies viral G gene sequences. In this study, we evaluated three methods, conventional RT-PCR and direct sequencing, adapter RT-PCR and sequencing with universal primers, conventional RT-PCR and cloning sequencing with universal primers, to detect rabies in animal brain homogenate. Four rabies isolates recovered from Fuyang city of Anhui province were diagnosed as positive using the fluorescentantibody test, rapid rabies enzyme immunodiagnosis methods and the mouse inoculation test. The results indicated that the adapter RT-PCR method and sequencing with universal primers is extremely well suited for sequencing rabies viral gene, which is rapid, cost-effective and precise compared with other two methods.<strong></strong></p>

scholarly journals Development of RT-PCR Based Diagnosis of SARS-CoV-2

2021 ◽  
Author(s):  
Rutuja Sunil Patankar ◽  
Vasudeo Pandharinath Zambare
Keyword(s):  
World Wide ◽  
Method Development ◽  
Vaccine Development ◽  
Whole Genome Sequence ◽  
Molecular Technique ◽  
Rt Pcr ◽  
Global Priority ◽  
Diagnosis Method ◽  
Pcr Method ◽  
Polymerase Chain

In the 2020, COVID-19 pandemic disease created an havoc situation world widely and mainly caused by Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2). It has been challenging task for researchers, scientists and medico-pharmaceutical organisations to find out rapid and reliable diagnosis methods. Among the all testing services, a Reverse Transcription Polymerase Chain Reaction (RT-PCR) is the more accurate, rapid and authenticated molecular technique used for most of the diagnosis of major diseases. It has been a global priority to fix the rapid diagnosis method to combat against the pandemic COVID-19. Thus, the present chapter mainly focussing on the progress of RT-PCR method development though various processes of data collection on isolation of whole genome sequence, its primer and method designing. In this scenario, India suddenly become the global leader for vaccine development and hence the challenges and RT-PCR kit development in India and rest of the world has been be discussed. World wide many Government and private agencies and industries have taken an initiative for diagnosis of SARS-CoV-2 hence this chapter also summarised the scope of RT-PCR to combat pandemic situation in future.

Cloning of GRK2 cDNA from S49 murine lymphoma cells

1996 ◽  
Vol 270 (3) ◽  
pp. C885-C891 ◽  
Author(s):  
R. J. Hughes ◽  
K. L. Anderson ◽  
D. Kiel ◽  
P. A. Insel
Keyword(s):  
Adrenergic Receptor ◽  
Direct Sequencing ◽  
Single Step ◽  
Receptor Kinase ◽  
Rt Pcr ◽  
Lymphoma Cells ◽  
Mouse Lymphoma Cells ◽  
Pcr Products ◽  
Polymerase Chain ◽  
Mouse Lymphoma

Beta-adrenergic receptor kinase is a member of the G protein-linked receptor kinase (GRK1) family that elicits receptor desensitization. We have cloned GRK2 from S49 mouse lymphoma cells. The nucleotide sequences of rat GRK2 and GRK3 were aligned and conserved primers chosen for use in reverse transcription-polymerase chain reaction (RT-PCR) of S49 mRNA. Direct sequencing of the PCR fragment provided a rapid means to identify the expression of the GRK2 but not the GRK3 transcript in these cells. Unique expression of GRK2 in S49 cells was confirmed by Western blotting. Three additional pairs of primers were chosen from the rat GRK2 sequence to amplify overlapping regions that together encompassed the entire coding sequence. After attempts to ligate the four fragments of S49 cell GRK2 cDNA by using PCR proved unsuccessful, the intact cDNA was assembled by digesting the PCR products in the region of the overlaps and ligating them in a single step into pBlue-script SK(+).

scholarly journals Mass scale screening of common arboviral infections by an affordable, cost effective RT–PCR method

2012 ◽  
Vol 2 (2) ◽  
pp. 97-101 ◽  
Author(s):  
Debjani Taraphdar ◽  
Arindam Sarkar ◽  
Shyamalendu Chatterjee
Keyword(s):  
Cost Effective ◽  
Mass Scale ◽  
Rt Pcr ◽  
Pcr Method

Immuno-capture PCR method for detecting Acidovorax avenae subsp. citrulli from watermelon

2007 ◽  
Vol 4 (2) ◽  
pp. 173-179 ◽  
Author(s):  
Wang Xiao ◽  
Zhang Le ◽  
Xu Fu-Shou ◽  
Zhao Li-Han ◽  
Xie Guan-Lin
Keyword(s):  
Polymerase Chain Reaction ◽  
Low Cost ◽  
Chain Reaction ◽  
Direct Pcr ◽  
Causal Organism ◽  
Bacterial Fruit Blotch ◽  
Pcr Method ◽  
Polymerase Chain ◽  
Honeydew Melon ◽  
Melon Seeds

AbstractAn immuno-capture polymerase chain reaction (IC-PCR) method for detection of Acidovorax avenae subsp. citrulli (AAC), the causal organism of bacterial fruit blotch (BFB) of watermelon, was developed by combining the immunosorbent enrichment (ISE) method with classical PCR and comparing with the direct PCR and growth check methods. The results showed that all A. avenae subsp. citrulli strains tested have produced 360 bp specific fragments using IC-PCR and direct PCR methods, while other strains from 10 different genera showed negative PCR results. The minimum detection concentration was about 50–100 cfu/ml and 104 cfu/ml, respectively. The IC-PCR sensitivity was 100 times higher than that of direct PCR. The examination of seven batches of different melon seeds from the markets by IC-PCR showed that one cantaloupe, two honeydew melon and two watermelon seed varieties carried the pathogen, indicating that the IC-PCR is an accurate, sensitive, rapid and low-cost technique.

scholarly journals A revisit to a low-cost method for the isolation of microsatellite markers: the case of the endangered Malayan tapir (Tapirus indicus)

2018 ◽  
Author(s):  
Qi Luan Lim ◽  
Nurul Adilah Ismail ◽  
Ramitha Arumugam ◽  
Wei Lun Ng ◽  
Christina Seok Yien Yong ◽  
...  
Keyword(s):  
Microsatellite Markers ◽  
Microsatellite Loci ◽  
Low Cost ◽  
Pcr Cloning ◽  
Chain Reaction ◽  
Sequencing Method ◽  
Rapid Polymerase Chain Reaction ◽  
Pcr Method ◽  
Polymerase Chain ◽  
Tapirus Indicus

AbstractThere are many approaches to develop microsatellite markers. We revisited an easy and rapid Polymerase Chain Reaction (PCR)-cloning-sequencing method to design microsatellite markers for Tapirus indicus. Using six random amplified microsatellite (RAM) markers, this study had rapidly generated 45 unique genomic sequences containing microsatellites. After screening 15 terminal and seven intermediate microsatellite loci, we shortlisted five and seven which were amplified either by single- or multiplex PCR using the economical three-primer PCR method. Genotyping attempts were made with ten Tapirus indicus individuals using three of the terminal microsatellite loci and all seven intermediate loci. However, none of the terminal microsatellite loci were considered useful for population genotyping studies, while the seven intermediate loci showed good amplification but were monomorphic in the ten samples. Despite successful detection of amplified loci, we would like to highlight that, researchers who are interested in this alternative method for isolation of microsatellite loci to be cautious and be aware of the limitations and downfalls reported herein that could render these loci unsuitable for population genotyping.

scholarly journals Deteksi Plasmodium falciparum dengan menggunakan metode real-time polymerase chain reaction di daerah Likupang dan Bitung

2016 ◽  
Vol 4 (1) ◽  
Author(s):  
David C. Tooy ◽  
Janno B. Bernadus ◽  
Angle Sorisi
Keyword(s):  
Polymerase Chain Reaction ◽  
Plasmodium Falciparum ◽  
Real Time ◽  
Real Time Pcr ◽  
18S Rrna ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Negative Results ◽  
Pcr Method ◽  
Polymerase Chain

Abstract: Malaria is one of the most important parasitic disease which is caused by Plasmodium spp. There are approximately 1,2 billion people in the world with high risk of getting malaria. Plasmodium falciparum (P. falciparum) is the cause of tropical malaria or falciparum malaria, and is responsible for most of the mortality rate. Currently, real-time polymerase chain reaction (RT-PCR) is being studied as an alterative of conventional malarian examination. Mangold et al reported that RT-PCR have 94.1% sensitivity and 100% specificity compared to microscopic examination in detecting P. falciparum. The aim of this research is to detect the presence of P. falciparum using RT-PCR in Likupang and Bitung region. This research were using descriptive design to find out the capability of real-time PCR method to detect P. falciparum in Likupang dan Bitung region. The researcher have examined 71 samples which are fulfill the research sample’s criteria. Postive results of P. falciparum found in 18 samples (25,3%) and negative results in 53 samples (74,6%) of total 71 samples with using RT-PCR. No positive results were found in samples from Likupang. There are positive result of P. falciparum in samples from Bitung. It is concluded that RT-PCR method can detect the presence of P. falciparum from the samples obtained from Likupang and Bitung based on the presence of its DNA. This detection efford is done by using 18S rRNA as target gene and ajust specific temperature on the RT-PCR instrument.Keywords: Plasmodium falciparum, Real-time Polymerase Chain Reaction (PCR), DetectionAbstrak: Malaria merupakan salah satu penyakit penting yang disebabkan oleh parasit Plasmodium spp. Kira-kira 1,2 miliar penduduk dunia memiliki risiko tinggi untuk mendapat malaria. Di Indonesia sendiri, terdapat 343.527 kasus terkonfirmasi dan 45 kematian karena malaria. Plasmodium falciparum (P. Falciparum) merupakan penyebab dari malaria tropika atau malaria falsiparum, dan bertanggung jawab atas sebagian besar angka mortalitas. Saat ini Real-Time Polymerase Chain Reaction (RT-PCR) telah banyak diteliti sebagai alternatif dari pemeriksaan malaria. Mangold dkk melaporkan bahwa real-time PCR memiliki nilai sensitivitas 94,1% dan nilai spesifisitas 100% terhadap pemeriksaan mikroskopis dalam mendeteksi P. falciparum. Penelitian bertujuan untuk mendeteksi P. falciparum dengan menggunakan RT-PCR di daerah Likupang dan Bitung. Penelitian ini menggunakan rancangan penelitian deskriptif untuk mengetahui kemampuan metode real-time PCR dalam mendeteksi P. falciparum di daerah Likupang dan Bitung. Tujuan penelitian ini ialah untuk mendeteksi keberadaan P. falciparum dengan menggunakan metode real-time PCR di daerah Likupang dan Bitung. Peneliti memeriksa 71 sampel darah yang memenuhi kriteria sampel penelitian. Hasil positif P. falciparum ditemukan pada 18 sampel (25,3 %) dan hasil negatif pada 53 sampel (74,6 %) dari total 71 sampel dengan menggunakan RT-PCR. Tidak ditemukannya hasil positif P. falciparum pada sampel dari Likupang. Ditemukan hasil positif P. falciparum pada sampel dari Bitung. Simpulan: Metode RT-PCR dapat mendeteksi P. falciparum berdasarkan keberadaan DNA-nya pada sampel yang diperoleh dari daerah Likupang dan Bitung. Deteksi ini berhasil dilakukan dengan menggunakan 18S rRNA sebagai gen target dan pengaturan suhu tertentu pada instrument RT-PCR.Kata kunci: P. falciparum, Real-time Polymerase Chain Reaction (PCR), Detection

scholarly journals Point-of-Care Diagnostic Tools for Surveillance of SARS-CoV-2 Infections

2021 ◽  
Vol 9 ◽  
Author(s):  
Dhanasekaran Sakthivel ◽  
David Delgado-Diaz ◽  
Laura McArthur ◽  
William Hopper ◽  
Jack S. Richards ◽  
...  
Keyword(s):  
Large Scale ◽  
Clinical Symptoms ◽  
Point Of Care ◽  
Cost Effective ◽  
Nucleic Acid Amplification ◽  
Diagnostic Tools ◽  
Rt Pcr ◽  
Serological Tests ◽  
Polymerase Chain ◽  
Effective Public Health

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is a recently emerged and highly contagious virus that causes coronavirus disease 2019 (COVID-19). As of August 24, 2021, there were more than 212 million confirmed COVID-19 cases and nearly 4.4 million deaths reported globally. Early diagnosis and isolation of infected individuals remains one of the most effective public health interventions to control SARS-CoV-2 spread and for effective clinical management of COVID-19 cases. Currently, SARS-CoV-2 infection is diagnosed presumptively based on clinical symptoms and confirmed by detecting the viral RNA in respiratory samples using reverse transcription polymerase chain reaction (RT-PCR). Standard RT-PCR protocols are time consuming, expensive, and technically demanding, which makes them a poor choice for large scale and point-of-care screening in resource-poor settings. Recently developed isothermal nucleic acid amplification tests (iNAAT), antigen and/or serological tests are cost-effective to scale COVID-19 testing at the point-of-care (PoC) and for surveillance activities. This review discusses the development of rapid PoC molecular tools for the detection and surveillance of SARS-CoV-2 infections.

scholarly journals COVID-19 Clinical Practice Guidelines Hadhramout Initiative Against Corona Group (HIAC Group)

2021 ◽  
Author(s):  
Abdulla A. Baradwan ◽  
Khalid H. Ba-Msahell ◽  
Iman A. Ba-Saddik ◽  
Ibrahim Albakri ◽  
Hassan O. Batis ◽  
...  
Keyword(s):  
Clinical Practice ◽  
Clinical Practice Guidelines ◽  
Practice Guidelines ◽  
Cost Effective ◽  
Case Definition ◽  
Rt Pcr ◽  
Care Workers ◽  
Chest X Ray ◽  
Polymerase Chain ◽  
Evidence Based Guidelines

Hadhramout Initiative Against Corona (HIAC ) formed a working group of clinicians relevant to the management of COVID-19 to formulate clinical practice guidelines (CPG) for clinicians caring for the patients infected with COVID-19 in Yemen. The regional guidelines on the management of COVID-19 were thoroughly reviewed and its applicability was assessed for Yemen. HIAC&rsquo;s recommendations covered (2) sections: the first one is the diagnosis, including case definition, risk stratification of the affected cases, investigations (prioritization of reverse transcription-polymerase chain reaction (RT-PCR) testing, D-dimer, chest x-ray, and chest computed tomography, while the second part covers the treatments (mainly Favipiravir, Remdesivir, Hydroxychloroquine, and Glucocorticoid, Anticoagulants, and Supportive measures). In conclusion, the adoption of cost-effective and evidence-based guidelines in Yemen will standardize the management of the patients infected with COVID-19 and protect both the patients and the health care workers from malpractice.

Sign in / Sign up

Export Citation Format

Share Document