Identification and Functional Analysis of a Defensin CcDef2 from Coridius chinensis
Abstract
:1. Introduction
2. Results
2.1. Characteristic Analysis of CcDef2
2.2. Multiple Sequence Alignment and Phylogenetic Analysis
2.3. Spatiotemporal Expression Profile
2.4. Expression Patterns of CcDef2 upon Bacterial Challenge
2.5. Expression Analysis of Recombinant Protein
2.6. Antibacterial Spectrum and MIC
3. Discussion
4. Materials and Methods
4.1. Insects and Bacteria
4.2. RNA Extraction and Gene Cloning
4.3. Bioinformatic Analyses
4.4. Spatiotemporal Expression Profile
4.5. Bacterial Infection Assay
4.6. Construction of Recombinant Expression Vector
4.7. Heterologous Expression, Purification and Refolding
4.8. Antimicrobial Assay
4.9. MIC Determination of CcDef2
4.10. Data Analysis
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Bacterial Strain | MIC (μM) |
---|---|
Gram-positive bacteria | |
S. aureus ATCC 25923 | 0.92 |
M. luteus CMCC 28001 | 1.24 |
B. subtilis CMCC 63501 | 1.56 |
Gram-negative bacteria | |
E. coli ATCC 25922 | ND |
P. aeruginosa CMCC 10104 | ND |
S. typhi CMCC 50071 | ND |
Primer Name | Sequence | Primer Usage |
---|---|---|
Def2-F | 5′-ACGCTTCAGTTGAGTCCATCT-3′ | Cloning |
Def2-R | 5′-ACAGTGATCTTTTGGTGTCCACT-3′ | |
Def2-qF | 5′-GCTGTCGCTGTTGTCTACATCGGT-3′ | RT-qPCR |
Def2-qR | 5′-CGGCTCTTCTTCGTGGTATGTCTC-3′ | |
Actin-F | 5′-ACCGCTGAGAGGGAAATCG-3′ | |
Actin-R | 5′-CAAGAAGGAAGGCTGGAAGAG-3′ |
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Gong, T.; Du, J.; Li, S.-W.; Huang, H.; Qi, X.-L. Identification and Functional Analysis of a Defensin CcDef2 from Coridius chinensis. Int. J. Mol. Sci. 2022, 23, 2789. https://doi.org/10.3390/ijms23052789
Gong T, Du J, Li S-W, Huang H, Qi X-L. Identification and Functional Analysis of a Defensin CcDef2 from Coridius chinensis. International Journal of Molecular Sciences. 2022; 23(5):2789. https://doi.org/10.3390/ijms23052789
Chicago/Turabian StyleGong, Tao, Juan Du, Shang-Wei Li, Hai Huang, and Xiao-Lang Qi. 2022. "Identification and Functional Analysis of a Defensin CcDef2 from Coridius chinensis" International Journal of Molecular Sciences 23, no. 5: 2789. https://doi.org/10.3390/ijms23052789