Development of PCR-Based Method for Rapid Detection of Abrin Gene
AbstractAbrin known as ribosome inactivating protein’s (RIP) it is a high cytotoxic plant protein. High lethality and low cost and easy access to plant and seeds lead to this toxin used in crimes and terrorist acts. Because obtaining purified toxins requires advanced laboratory equipment and complex procedures, so, it appears that the perpetrators of such crimes use crude extracts. As a result, remaining of specific toxin gene in these extracts take the advantage of using PCR assay to identify abrin gene, that refers to exist of its toxin. We use new rapid molecular method for abrin gene detection by PCR. We designed specific primers for detection of abrin toxins gene by PCR. DNA was extracted by cetyl trimethylammonium bromide-polyvinylpyrrolidone (CTAB-PVP) method from Rosary pie samples and PCR protocol was performed using primers for toxin gene amplification. Then we analyzed assay’s sensitivity by serial dilution method. The results of this study revealed that designed and selected primers sequence for toxin’s gene is specific. The desired Product size was obtained and Sequencing of PCR products was performed that show up to 90% similarity with known sequence for each molecule. According to these results, the developed rapid molecular method for detection of abrin toxin gene is sensitive, and low-cost to detect this toxins gene in cases of suspected to bioterrorism event.
2. Bradberry SM, Dickers KJ, Rice P, Griffiths GD, Vale JA. Ricin poisoning. Toxicological reviews. 2003;22(1):65-70.
3. Dickers KJ, Bradberry SM, Rice P, Griffiths GD, Vale JA. Abrin poisoning. Toxicological reviews. 2003;22(3):137-42.
4. Olsnes S, Sandvig K, Eiklid K, Pihl A. Properties and action mechanism of the toxic lectin modeccin: interaction with cell lines resistant to modeccin, abrin, and ricin. Journal of supramolecular structure. 1978;9(1):15-25.
5. Audi J, Belson M, Patel M, Schier J, Osterloh J. Ricin poisoning: a comprehensive review. Jama. 2005;294(18):2342-51.
6. Stirpe F, Battelli MG. Ribosome-inactivating proteins: progress and problems. Cellular and molecular life sciences : CMLS. 2006;63(16):1850-66.
7. Olsnes S, Refsnes K, Christensen TB, Pihl A. Studies on the structure and properties of the lectins from Abrus precatorius and Ricinus communis. Biochimica et biophysica acta. 1975;405(1):1-10.
8. Robertus J. The structure and action of ricin, a cytotoxic N-glycosidase. Seminars in cell biology. 1991;2(1):23-30.
9. Hutchkinson M. The Poisoner's Handbook: Loompanics Unlimited; 1988.
10. Integrated DNA Technologies. Plant DNA Extraction Protocol - Integrated DNA Technologies [cited 2016 22 August]. Available from: https://eu.idtdna.com/pages/docs/educational-resources/plant-dna-extraction-protocol.pdf?sfvrsn=5.
11. National Center for Biotechnology Information: Ncbi.nlm.nih.gov; [cited 2016 10/31]. Available from: https://www.ncbi.nlm.nih.gov/.
12. Untergasser A, Cutcutache I, Koressaar T, Ye J, Faircloth BC, Remm M, et al. Primer3--new capabilities and interfaces. Nucleic acids research. 2012;40(15):e115.
13. Integrated DNA Technologies. OligoAnalyzer 3.1.
14. Staroscik A. Calculator for determining the number
of copies of a template 2004 [updated 29 January 2004; cited 2016 23 August]. Available from: http://cels.uri.edu/gsc/cndna.html.
15. Bozza WP, Tolleson WH, Rosado LA, Zhang B. Ricin detection: tracking active toxin. Biotechnology advances. 2015;33(1):117-23.
16. Mayor S. UK doctors warned after ricin poison found in police raid. Bmj. 2003;326(7381):126.
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