LAMP Coupled CRISPR-Cas12a Module for Rapid, Sensitive and Visual Detection of Porcine Circovirus 2
Abstract
:Simple Summary
Abstract
1. Introduction
2. Materials and Methods
2.1. Viruses and Clinical Samples
2.2. Nucleic Acid Preparation
2.3. LAMP Primers Design and crRNA Preparation
2.4. Preparation of huLbCas12a
2.5. Determination of Target Cleavage Activity of the huLbCas12a Protein
2.6. Optimization and Evaluation of the LAMP-CRISPR Detection Method
2.7. LAMP-CRISPR Fluorescence Assay
2.8. Efficiency Assessment of the LAMP-CRISPR Detection System for Clinical Samples Detection
2.9. Statistic Analysis
3. Results
3.1. Expression and Purification of the huLbCas12a Protein in E. coli
3.2. Target Cleavage Activity of huLbCas12a Protein
3.3. Optimization of the LAMP-CRISPR Detection
3.4. Sensitivity and Specificity Test of LAMP-CRISPR Detection
3.5. Comparation of LAMP-CRISPR and Taqman-Based qPCR in Detecting Clinical Samples
4. Discussion
5. Conclusions
6. Patents
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Simple Number | 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 | 9 | 10 |
Ct value | - | 33.82 | 23.54 | 31.55 | 32.84 | 32.13 | - | - | 25.59 | 35.44 |
Simple number | 11 | 12 | 13 | 14 | 15 | 16 | 17 | 18 | 19 | 20 |
Ct value | 35.28 | 31.77 | 28.44 | 34.38 | 35.80 | 28.39 | 20.15 | - | - | 25.81 |
Simple number | 21 | 22 | 23 | 24 | 25 | 26 | 27 | 28 | 29 | 30 |
Ct value | 25.35 | - | - | - | 17.14 | 31.19 | 35.02 | - | - | 32.71 |
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Lei, L.; Liao, F.; Tan, L.; Duan, D.; Zhan, Y.; Wang, N.; Wang, Y.; Peng, X.; Wang, K.; Huang, X.; et al. LAMP Coupled CRISPR-Cas12a Module for Rapid, Sensitive and Visual Detection of Porcine Circovirus 2. Animals 2022, 12, 2413. https://doi.org/10.3390/ani12182413
Lei L, Liao F, Tan L, Duan D, Zhan Y, Wang N, Wang Y, Peng X, Wang K, Huang X, et al. LAMP Coupled CRISPR-Cas12a Module for Rapid, Sensitive and Visual Detection of Porcine Circovirus 2. Animals. 2022; 12(18):2413. https://doi.org/10.3390/ani12182413
Chicago/Turabian StyleLei, Lei, Fan Liao, Lei Tan, Deyong Duan, Yang Zhan, Naidong Wang, Yuge Wang, Xiaoye Peng, Kaixin Wang, Xiaojiu Huang, and et al. 2022. "LAMP Coupled CRISPR-Cas12a Module for Rapid, Sensitive and Visual Detection of Porcine Circovirus 2" Animals 12, no. 18: 2413. https://doi.org/10.3390/ani12182413
APA StyleLei, L., Liao, F., Tan, L., Duan, D., Zhan, Y., Wang, N., Wang, Y., Peng, X., Wang, K., Huang, X., Yang, Y., & Wang, A. (2022). LAMP Coupled CRISPR-Cas12a Module for Rapid, Sensitive and Visual Detection of Porcine Circovirus 2. Animals, 12(18), 2413. https://doi.org/10.3390/ani12182413