Designing Two Synthetic Constructs for Real Time PCR Detection of Francisella tularensis and Ebola Virus.

Content Provider	Europe PMC
Author	Dehghan Esmat Abadi, Mohammadjavad Motalebzadeh, Hesam Barati, Mahmoud Yaghobi, Mohammadali
Copyright Year	2021
Abstract	Background:Generally, timely diagnosis of micro-organisms is very important to prevent many diseases. Many methods can detect micro-organisms like culture-based methods and molecular methods. The molecular methods are usually preferred because they provide fast and reliable results. In some cases, microbial strains are not accessible, and there is no safety to work with them; therefore, synthetic constructs which are designed according to the available sequences in databases can be used as a positive control for detection of them.Methods:In this study, a synthetic construct was designed for molecular detection of Francisella tularensis (F. tularensis) and the Ebola virus by multiplex real-time PCR reaction. For this, sequences were taken from databases and then multiple alignments were done by software. Also, conventional PCR and two models of real-time PCR (SYBR green and TaqMan) were applied. Finally, multiplex real-time PCR was performed.Results:The synthetic construct was designed and used for conventional PCR and multiplex PCR. The results of common PCR showed a single band at 148 bp and 167 bp in 1.5% agarose gel stained by ethidium bromide for F. tularensis and Ebola virus, respectively. Also, a dual-band at 148 and 167 bp was observed in multiplex PCR. Results of real-time PCR showed a limit of detection about 0.1 pg of plasmid/μl.Conclusion:In conclusion, the designed construct can be used as a positive control for an accurate diagnosis of these micro-organisms without any biological danger for laboratory staff. So, this method is useful for diagnosis of these agents in food, water, and blood samples.
Related Links	https://europepmc.org/backend/ptpmcrender.fcgi?accid=PMC8112141&blobtype=pdf
ISSN	20082835
Journal	Avicenna Journal of Medical Biotechnology [Avicenna J Med Biotechnol]
Volume Number	13
DOI	10.18502/ajmb.v13i2.5528
PubMed Central reference number	PMC8112141
Issue Number	2
PubMed reference number	34012525
e-ISSN	20084625
Language	English
Publisher	Avicenna Research Institute
Publisher Date	2021-04-01
Access Restriction	Open
Rights License	This is an open access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited, appropriate credit is given, any changes made indicated, and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/ Copyright© 2021 Avicenna Research Institute
Subject Keyword	Ebola virus Humans Multiplex polymerase chain reaction Real-time polymerase chain reaction
Content Type	Text
Resource Type	Article
Subject	Bioengineering Biomedical Engineering Applied Microbiology and Biotechnology Biotechnology