An integrated digital PCR system with high universality and low cost for nucleic acid detection.

Kangning Wang, Bin Li, Yu Guo, Yanqi Wu, Yan Li, Wenming Wu
Author Information
  1. Kangning Wang: Institute of Biological and Medical Engineering, Guangdong Academy of Sciences, Guangzhou, China.
  2. Bin Li: Institute of Microbiology Chinese Academy of Sciences, Beijing, China.
  3. Yu Guo: School of Mechanical and Electrical Engineering, Guangdong University of Technology, Guangzhou, China.
  4. Yanqi Wu: State Key Laboratory of Quality Research in Chinese Medicine, Macau University of Science and Technology, Taipa, China.
  5. Yan Li: Institute of Biological and Medical Engineering, Guangdong Academy of Sciences, Guangzhou, China.
  6. Wenming Wu: Institute of Biological and Medical Engineering, Guangdong Academy of Sciences, Guangzhou, China.

Abstract

Digital PCR is the most advanced PCR technology. However, due to the high price of the digital PCR analysis instrument, this powerful nucleic acid detection technology is still difficult to be popularized in the general biochemistry laboratory. Moreover, one of the biggest disadvantages of commercial digital PCR systems is the poor versatility of reagents: each instrument can only be used for a few customized kits. Herein, we built a low-cost digital PCR system. The system only relies on low-cost traditional flat-panel PCR equipment to provide temperature conditions for commercial dPCR chips, and the self-made fluorescence detection system is designed and optically optimized to meet a wide range of reagent requirements. More importantly, our system not only has a low cost (<8000 US dollars) but also has a much higher universality for nucleic acid detection reagents than the traditional commercial digital PCR system. In this study, several samples were tested. The genes used in the experiment were plasmids containing UPE-1a fragment, TP53 reference DNA, hepatitis B virus DNA, leukemia sample, SARS-COV-2 DNA, and SARS-COV-2 RNA. Under the condition that DNA can be amplified normally, the function of the dPCR system can be realized with simpler and low-price equipment. Some DNA cannot be detected by using the commercial dPCR system because of the special formula when it is configured as the reaction solution, but these DNA fluorescence signals can be clearly detected by our system, and the concentration can be calculated. Our system is more applicable than the commercial dPCR system to form a new dPCR system that is smaller and more widely applicable than commercially available machinery.

Keywords

References

  1. Anal Chem. 2012 Jan 17;84(2):1003-11 [PMID: 22122760]
  2. Biosens Bioelectron. 2018 Nov 30;120:144-152 [PMID: 30173010]
  3. Clin Exp Ophthalmol. 2019 May;47(4):513-520 [PMID: 30414235]
  4. JAMA Oncol. 2016 Aug 1;2(8):1014-22 [PMID: 27055085]
  5. Lab Chip. 2016 Jan 7;16(1):208-16 [PMID: 26610263]
  6. Ann Oncol. 2017 Jul 01;28(7):1508-1516 [PMID: 28472366]
  7. PLoS One. 2011 Mar 08;6(3):e17791 [PMID: 21408138]
  8. Nat Methods. 2009 Jul;6(7):503-6 [PMID: 19525960]
  9. Clin Biochem. 2015 Oct;48(15):948-56 [PMID: 25828047]
  10. Light Sci Appl. 2013;2: [PMID: 24527266]
  11. Dig Dis Sci. 2012 Apr;57(4):897-904 [PMID: 22198701]
  12. Clin Chem. 2013 Dec;59(12):1732-41 [PMID: 24061615]
  13. Exp Hematol. 2002 Jun;30(6):503-12 [PMID: 12063017]
  14. Expert Rev Mol Diagn. 2005 Mar;5(2):209-19 [PMID: 15833050]
  15. Lab Chip. 2018 Dec 4;18(24):3717-3732 [PMID: 30402632]
  16. Anal Chem. 2011 Sep 1;83(17):6474-84 [PMID: 21446772]
  17. BMC Cancer. 2017 Jun 19;17(1):428 [PMID: 28629339]
  18. BMC Mol Biol. 2008 May 06;9:46 [PMID: 18460208]
  19. Lancet Respir Med. 2018 Sep;6(9):681-690 [PMID: 30017884]
  20. Acta Trop. 2013 Jan;125(1):23-31 [PMID: 22982466]
  21. Analyst. 2019 Mar 25;144(7):2239-2247 [PMID: 30663740]
  22. Nucleic Acids Res. 2012 Jun;40(11):e82 [PMID: 22373922]

Word Cloud

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