scholarly journals Selective detection of Pseudomonas syringae pv. tomato using dot blot hybridization and real-time PCR

2007 ◽  
Vol 56 (4) ◽  
pp. 683-691 ◽  
Author(s):  
V. Fanelli ◽  
C. Cariddi ◽  
M. Finetti-Sialer
Keyword(s):  
Real Time ◽  
Pseudomonas Syringae ◽  
Real Time Pcr ◽  
Blot Hybridization ◽  
Selective Detection ◽  
Dot Blot ◽  
Dot Blot Hybridization

scholarly journals Comparison of a one-step real-time RT-PCR and a nested real-time RT-PCR for a genogroup II norovirus reveals differences in sensitivity depending upon assay design and visualization

PLoS ONE ◽  
2021 ◽  
Vol 16 (4) ◽  
pp. e0248581
Author(s):  
Clyde S. Manuel ◽  
Cassandra Suther ◽  
Matthew D. Moore ◽  
Lee-Ann Jaykus
Keyword(s):  
Detection Limit ◽  
Real Time ◽  
Real Time Pcr ◽  
Environmental Samples ◽  
Blot Hybridization ◽  
Molecular Techniques ◽  
Rt Pcr ◽  
Dot Blot ◽  
Dot Blot Hybridization ◽  
One Step

Human norovirus (NoV) is the leading cause of acute viral gastroenteritis and a major source of foodborne illness. Detection of NoV in food and environmental samples is typically performed using molecular techniques, including real-time reverse transcription polymerase chain reaction (RT-PCR) and less frequently, nested real-time PCR. In this study, we conducted a controlled comparison of two published NoV detection assays: a broadly reactive one-step real-time RT-PCR and a two-step nested real-time PCR assay. A 20% human fecal suspension containing a genogroup II human NoV was serially diluted, genome extracted, and subjected to amplification using the two assays compared via PCR Units. Additional amplicon confirmation was performed by dot blot hybridization using digoxigenin (DIG)-labeled oligonucleotide probes. Both assays displayed similar amplification standard curves/amplification efficiencies; however, the nested assay consistently detected one log10 lower virus. Dot blot hybridization improved the detection limit of the nested real-time PCR by one log10 NoV genome copies but impaired the detection limit of the one-step real-time RT-PCR by one log10 NoV genome copies. These results illustrate the complexities in designing and interpreting molecular techniques having a sufficient detection limit to detect low levels of viruses that might be anticipated in contaminated food and environmental samples.

scholarly journals An Integrated Diagnostic Approach for Citrus Exocortis Viroid

2021 ◽  
Author(s):  
Chih-Hsu Lin ◽  
Ting-Hsuan Hung ◽  
I Hu ◽  
Ta-Hsin Ku ◽  
Chun-Yi Lin ◽  
...  
Keyword(s):  
Real Time ◽  
Reverse Transcription ◽  
Severe Disease ◽  
Blot Hybridization ◽  
Rt Pcr ◽  
Dot Blot ◽  
Tomato Cultivar ◽  
Dot Blot Hybridization ◽  
Reverse Transcription Pcr ◽  
One Step

Abstract BackgroundCitrus exocortis viroid (CEVd) is a circular single-stranded RNA pathogen consists of around 370 nucleotides and leads to a severe disease showing bark scaling symptom on citrus crops, which leads to yield decrease and economic loss. Since the absence of viroid-encoded proteins, methods for CEVd detection mainly counts on bioassays or nucleic acid-base approaches. In order to validate the CEVd disease, here we developed an integrated diagnostic protocol. MethodsCEVd transcripts were inoculated onto two susceptible cultivars of Solanum lycopersicum L., cv. Rutgers and cv. Double-Fortune, seedings. After inoculation, total RNAs of the two tomato cultivars were extracted to detect CEVd infection by dot blot hybridization, one-step reverse transcription PCR (one-step RT-PCR) and real-time reverse transcription PCR (real-time RT-PCR). In addition, the symptom development of both cultivars was recorded weekly. ResultsThe tomato cultivar Rutgers rather than Double-Fortune or others was selected as a suitable CEVd-indicator plant and the bio-index score was established based on epinasty, vein necrosis, leaf size reduction and stunting symptoms. In addition, the isolate of CEVd that collected from citrus field could rapidly and consistently cause the index symptoms on Rutgers. As expected, CEVd could be specifically and sensitively detected in both tomato and citrus plants by dot-blot hybridization and RT-PCR technologies, including one-step RT-PCR and real-time RT-PCR. Furthermore, we found that the levels of CEVd genomic RNA or CEVd derived small RNAs are correlated to symptom severity. ConclusionsIn this study, we developed an integrated detection method for CEVd and revealed potential underlying viroid-host interactions.

APPLICATION OF REAL-TIME PRC TECHNIQUE AND REVERSE DOT-BLOT FOR DETECTION THE HIGH RISK TYPES OF HUMAN PAPILLOMAVIRUS CAUSING CERVICAL CANCER

2017 ◽  
pp. 99-103
Author(s):  
Van Bao Thang Phan ◽  
Hoang Bach Nguyen ◽  
Van Thanh Nguyen ◽  
Thi Nhu Hoa Tran ◽  
Viet Quynh Tram Ngo
Keyword(s):  
Cervical Cancer ◽  
High Risk ◽  
Real Time ◽  
Real Time Pcr ◽  
Dot Blot ◽  
The Real ◽  
Dot Blot Assay ◽  
Hpv Types ◽  
High Risk Hpv ◽  
Reverse Dot Blot

Introduction: Infection with HPV is the main cause of cervical cancer. Determining HPV infection and the types of HPV plays an important role in diagnosis, treatment and prognosis of cervicitis/cervical cancer. Aims: Determining proportion of high-risk HPV types and the occurrence of coinfection with multiple HPV types. Methods: 177 women with cervicitis or abnormal Pap smear result were enrolled in the study. Performing the real-time PCR for detecting HPV and the reverse DOT-BLOT assay for determining type of HPV in cases of positive PCR. Results: 7 types of high-risk HPV was dectected, the majority of these types were HPV type 18 (74.6%) and HPV type 16 (37.6%); the proportion of infection with only one type of HPV was 30.4% and coinfection with multiple HPV types was higher (69.6%), the coinfected cases with 2 and 3 types were dominated (32.2% and 20.3%, respectively) and the coinfected cases with 4 and 5 types were rare. Conclusion: Use of the real-time PCR and reverse DOT-BLOT assay can determine the high-risk HPV types and the occurrence of coinfection with multiple HPV types. Key words: HPV type, Reverse DOT-BLOT, real-time PCR,PCR, cervical cancer

Feasibility of Qualitative Testing of BCR-ABL and JAK2 V617F in Suspected Myeloproliperative Neoplasm (MPN) Using RT-PCR Reversed Dot Blot Hybridization (RT-PCR RDB)

2019 ◽  
Vol 19 (4) ◽  
pp. 220-227
Author(s):  
Najmiatul Masykura ◽  
Ummu Habibah ◽  
Siti Fatimah Selasih ◽  
Soegiarto Gani ◽  
Cosphiadi Irawan ◽  
...  
Keyword(s):  
Blot Hybridization ◽  
Jak2 V617f ◽  
Rt Pcr ◽  
Dot Blot ◽  
Dot Blot Hybridization

Prevalence of HPV in a Melbourne Female STD Population: Comparison of RNA and DNA Probes in Detecting HPV by Dot Blot Hybridization

1993 ◽  
Vol 4 (3) ◽  
pp. 159-164 ◽  
Author(s):  
A J Borg ◽  
G Medley ◽  
S M Garland
Keyword(s):  
Past History ◽  
Blot Hybridization ◽  
Condylomata Acuminata ◽  
Dot Blot ◽  
Hpv Dna ◽  
Sexually Transmitted ◽  
Dot Blot Hybridization ◽  
History Of ◽  
Cytological Features ◽  
Dna Types

A total of 377 women, consecutively selected as first attenders to a sexually transmitted diseases clinic in Melbourne, Australia, were examined for overt Condylomata acuminata and were screened for genital HPV DNA types 6, 11, 16, 18, 31, 33 and (35) using 2 dot blot hybridization methods. Overall, there was a 90% positivity correlation between the 2 methods with HPV DNA being detected in 12% of ectocervical samples. Overt warts were found in 15% of the women and HPV DNA was detected at the cervix in 35% with cytology predicting HPV with or without dysplasia in 27%. Thirteen percent had a past history of warts but none on examination and HPV DNA was evident in 16% while 18% had cytological features of HPV. Those with no warts evident and no past history of warts had both HPV DNA and cytological features of HPV in 7%.

Detection of herpes simplex virus and adenovirus DNA by dot blot hybridization using in vitro synthesized RNA transcripts

1986 ◽  
Vol 13 (4) ◽  
pp. 291-299 ◽  
Author(s):  
Volker Schuster ◽  
Bertfried Matz ◽  
Helga Wiegand ◽  
Brigitte Traub ◽  
Dieter Neumann-Haefelin
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Blot Hybridization ◽  
Dot Blot ◽  
Rna Transcripts ◽  
Dot Blot Hybridization ◽  
Simplex Virus

Detection of Cryptosporidium parvum Oocysts on Fresh Vegetables and Herbs Using Antibodies Specific for a Cryptosporidium parvum Viral Antigen

2005 ◽  
Vol 68 (5) ◽  
pp. 1093-1096 ◽  
Author(s):  
K. E. KNIEL ◽  
M. C. JENKINS
Keyword(s):  
Cryptosporidium Parvum ◽  
Viral Antigen ◽  
Blot Hybridization ◽  
Surface Antigens ◽  
Food Samples ◽  
Sensitive Detection ◽  
Dot Blot ◽  
Dot Blot Hybridization ◽  
Fresh Vegetables ◽  
Cryptosporidium Parvum Oocysts

The purpose of this study was to determine if the viral symbiont of Cryptosporidium parvum (CPV) sporozoites could be used as a target for sensitive detection of the parasite in food samples. Polyclonal sera specific to the recombinant viral capsid protein (rCPV40) was used in a dot blot hybridization assay to detect oocysts recovered from green onions and cilantro. Small batches of chopped green onions and cilantro leaves were artificially contaminated with three different concentrations of oocysts: 106, 102, and 101. rCPV40 was superior in detecting oocysts compared with other antibodies directed toward total oocyst protein and oocyst surface antigens. This study provides evidence that CPV is an excellent target for sensitive detection of C. parvum oocysts in foods.

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