scholarly journals Real-Time Detection of the Mutation Responsible for Progressive Rod-Cone Degeneration in Labrador Retriever Dogs Using Locked Nucleic Acid TaqMan Probes

2009 ◽  
Vol 21 (5) ◽  
pp. 689-692 ◽  
Author(s):  
Fabio Gentilini ◽  
Gian Luca Rovesti ◽  
Maria Elena Turba
Keyword(s):  
Nucleic Acid ◽  
Real Time ◽  
Real Time Pcr ◽  
Late Onset ◽  
Labrador Retriever ◽  
Locked Nucleic Acid ◽  
Sensitivity Threshold ◽  
Ophthalmological Examination ◽  
Pcr Assay ◽  
Labrador Retrievers

Progressive rod-cone degeneration ( prcd) is a late onset, autosomal recessive, inherited disease in dogs caused by a G > A substitution in the PRCD locus, which has been reported in more than 18 breeds, including Labrador Retriever dogs. In this study, a real-time polymerase chain reaction (PCR) assay, exploiting the features of locked nucleic acid (LNA) fluorescent-labeled probes, was developed to genotype the sequence variants responsible for the disease. Two Labrador Retrievers were diagnosed with prcd by ophthalmological examination performed by a panelist of the Italian hereditary eye disease control program. The 2 dogs, as well as 8 related and 14 unrelated Labrador Retrievers, were genotyped with both direct sequencing of the disease locus and real-time LNA TaqMan PCR assay. Even though the region surrounding the mutation was predicted to be highly structured, making probe annealing difficult, the real-time PCR assay allowed researchers to correctly genotype the dogs in all cases with a sensitivity threshold of 4 ng/reaction of genomic DNA. A real-time PCR assay will allow a high-throughput analysis of a larger cohort of dogs, thereby enabling researchers to investigate the prevalence of the mutated allele in the affected breeds.

scholarly journals Locked Nucleic Acid Probe-Based Real-Time PCR Assay for the Rapid Detection of Rifampin-Resistant Mycobacterium tuberculosis

PLoS ONE ◽  
2015 ◽  
Vol 10 (11) ◽  
pp. e0143444 ◽  
Author(s):  
Yong Zhao ◽  
Guilian Li ◽  
Chongyun Sun ◽  
Chao Li ◽  
Xiaochen Wang ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Nucleic Acid ◽  
Real Time ◽  
Real Time Pcr ◽  
Rapid Detection ◽  
Locked Nucleic Acid ◽  
Pcr Assay ◽  
Locked Nucleic Acid Probe

scholarly journals New Real-Time PCR Assay Using Locked Nucleic Acid Probes To Assess Prevalence of ParC Mutations in Fluoroquinolone-Susceptible Streptococcus pneumoniae Isolates from France

2006 ◽  
Vol 50 (4) ◽  
pp. 1594-1598 ◽  
Author(s):  
Jean-Winoc Decousser ◽  
Imen Methlouthi ◽  
Patrick Pina ◽  
Anne Collignon ◽  
Pierre Allouch
Keyword(s):  
Streptococcus Pneumoniae ◽  
Nucleic Acid ◽  
High Risk ◽  
Real Time ◽  
Real Time Pcr ◽  
Locked Nucleic Acid ◽  
Pcr Assay ◽  
Nucleic Acid Probes ◽  
National Surveys ◽  
Risk Patients

ABSTRACT A real-time PCR assay with locked nucleic acid probes was developed to screen mutations at codons 79 and 83 of the Streptococcus pneumoniae parC gene. Only silent mutations were detected among 236 French invasive fluoroquinolone-susceptible strains. This test could be useful for some high-risk patients or in national surveys.

scholarly journals Differing Effects of Standard and Harsh Nucleic Acid Extraction Procedures on Diagnostic Helminth Real-Time PCRs Applied to Human Stool Samples

Pathogens ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 188
Author(s):  
Tanja Hoffmann ◽  
Andreas Hahn ◽  
Jaco J. Verweij ◽  
Gérard Leboulle ◽  
Olfert Landt ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Real Time ◽  
Real Time Pcr ◽  
Acid Extraction ◽  
Nucleic Acid Extraction ◽  
Pcr Assay ◽  
Bead Beating ◽  
Stool Samples ◽  
Pcr Assays ◽  
Human Stool

This study aimed to assess standard and harsher nucleic acid extraction schemes for diagnostic helminth real-time PCR approaches from stool samples. A standard procedure for nucleic acid extraction from stool and a procedure including bead-beating as well as proteinase K digestion were compared with group-, genus-, and species-specific real-time PCR assays targeting helminths and nonhelminth pathogens in human stool samples. From 25 different in-house and commercial helminth real-time PCR assays applied to 77 stool samples comprising 67 historic samples and 10 external quality assessment scheme samples positively tested for helminths, higher numbers of positive test results were observed after bead-beating-based nucleic acid extraction for 5/25 (20%) real-time PCR assays irrespective of specificity issues. Lower cycle threshold values were observed for one real-time PCR assay after the standard extraction scheme, and for four assays after the bead-beating-based scheme. Agreement between real-time PCR results after both nucleic acid extraction strategies according to Cohen’s kappa ranged from poor to almost perfect for the different assays. Varying agreement was observed in eight nonhelminth real-time PCR assays applied to 67 historic stool samples. The study indicates highly variable effects of harsh nucleic acid extraction approaches depending on the real-time PCR assay used.

Molecular Detection of Adenovirus in Tap Water from Coastal Areas Of Karachi Pakistan:The Real-Time PCR Assay

2021 ◽  
Vol 156 (Supplement_1) ◽  
pp. S140-S140
Author(s):  
A Kalam
Keyword(s):  
Nucleic Acid ◽  
Real Time ◽  
Low Income ◽  
Water Samples ◽  
Real Time Pcr ◽  
Tap Water ◽  
Watery Diarrhea ◽  
Acid Extraction ◽  
Pcr Assay ◽  
Viral Contamination

Abstract Introduction/Objective Diarrhea is a major source of morbidity and mortality in low-income and middle-income countries. In underdeveloped countries, diseases caused by viruses identified in environmental samples cause major health problems. Little knowledge about the frequency and pattern of viral contamination of drinking water sources in these resource-poor settings. Adenovirus which causes watery diarrhea, particular has been recognized as important causal pathogen. Adenovirus remains a global threat to public health and an indicator of inequity and lack of social development. Tap water samples from coastal sites in Karachi between 2019 and 2020 over a period of 11 months. The total of 40 tap water sample was examined for infectious Adenovirus by a real time polymerase chain reaction (PCR) amplification. Methods/Case Report This Pilot study is conducted on tap water samples from Karachi Pakistan, n=40 are processed. Extraction of nucleic acid from all filtered water samples collected with Sterivex filter units by using Qiagen DNeasy Power Water Sterivex Kit. As per the manufacturer’s instruction. Phocine herpesvirus(PhHV) is added as an external positive control to monitor the efficiency of nucleic acid extraction and amplification. TaqMan Universal PCR Master Mix (Thermo Fisher Scientific) is being used in probe based real time PCR assay,the below 35 Ct value is considered as a positive sample. Results (if a Case Study enter NA) Results showed the total of 37.7% of the sources were positive for adenovirus.The level of viral contamination was moderate to high. Conclusion The results has been showed that no seasonal pattern for viral contaminations was found after samples obtained during the dry and wet seasons were compared. Further the Real time PCR assay increases the sensitivity and provides the high resolution of pathogen detection.

scholarly journals Locked nucleic acid inhibits amplification of contaminating DNA in real-time PCR

BioTechniques ◽  
2005 ◽  
Vol 38 (4) ◽  
pp. 605-610 ◽  
Author(s):  
Lone Hummelshoj ◽  
Lars P. Ryder ◽  
Hans O. Madsen ◽  
Lars K. Poulsen
Keyword(s):  
Nucleic Acid ◽  
Real Time ◽  
Real Time Pcr ◽  
Locked Nucleic Acid

scholarly journals Detection of Spinal Muscular Atrophy Using a Duplexed Real-Time PCR Approach With Locked Nucleic Acid-Modified Primers

2021 ◽  
Vol 41 (1) ◽  
pp. 101-107
Author(s):  
Jianyan Pan ◽  
Chunhua Zhang ◽  
Yanling Teng ◽  
Sijing Zeng ◽  
Siyi Chen ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Spinal Muscular Atrophy ◽  
Real Time ◽  
Real Time Pcr ◽  
Muscular Atrophy ◽  
Locked Nucleic Acid
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