In order to explore the feasibility of the new rodenticide of type D Botulinum toxin to prevent and control the population density of pests in farmland from a molecular perspective. This study aimed to analyze the feasibility of D-type botulinum toxin for the control of urban rodents from a molecular perspective, and to detect the cleavage effect of D-type botulinum neurotoxin light chain recombinant protein on the synaptic vesicle membrane protein VAMP-1 of brown house mouse. The genomic DNA of Clostridium botulinum D 8901 strain was used as a template. Based on the botulinum toxin D gene sequence reported in GenBank, specific primers were designed. Plasmids were constructed by gene synthesis and subcloned into the pET28a expression vector. The recombinant plasmid was transformed into BL21 (DE3) Rosetta competent cells, and induced expression by IPTG. The expression products were purified by Ni-NTA affinity chromatography, identified by SDS-PAGE and Western blotting, and the recombinant proteins were analyzed by SDS-PAGE and Western blotting. The cleavage of VAMP1 protein in SD brown house mouse. The pET-28a- BDLc expression plasmid was successfully constructed and transformed into E. coli BL21 (DE3) Rosetta. Western blot confirmed that the recombinant protein pET-28a- BDLc (residues Met 1-Met 94) obtained soluble expression, and obtained 5 mg, Recombinant protein with purity > 90%. It has good biological activity as determined by animal method. The recombinant protein can decompose VAMP1 protein into two fragments. Type D botulinum toxin protein can specifically cleave synaptic vesicle membrane protein (VAMP1) of brown house mouse, and type D botulinum toxin is feasible for urban rodent control.
| Published in | Animal and Veterinary Sciences (Volume 12, Issue 6) |
| DOI | 10.11648/j.avs.20241206.14 |
| Page(s) | 171-176 |
| Creative Commons |
This is an Open Access article, distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution and reproduction in any medium or format, provided the original work is properly cited. |
| Copyright |
Copyright © The Author(s), 2024. Published by Science Publishing Group |
Botulinum Neurotoxin Type D, Expression of Recombinant Protein, Detection of Biological Activity
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APA Style
Shengqing, L., Shengyi, H., Shuping, L., Guoyuan, H., Huaixing, L., et al. (2024). Construction of Recombinant Protein of Botulinum Neurotoxin Light Chain and Analysis of Cleavage Effect on VAMP1 Protein in Brown Rat. Animal and Veterinary Sciences, 12(6), 171-176. https://doi.org/10.11648/j.avs.20241206.14
ACS Style
Shengqing, L.; Shengyi, H.; Shuping, L.; Guoyuan, H.; Huaixing, L., et al. Construction of Recombinant Protein of Botulinum Neurotoxin Light Chain and Analysis of Cleavage Effect on VAMP1 Protein in Brown Rat. Anim. Vet. Sci. 2024, 12(6), 171-176. doi: 10.11648/j.avs.20241206.14
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Download@article{10.11648/j.avs.20241206.14,
author = {Li Shengqing and Han Shengyi and Li Shuping and Hu Guoyuan and Liu Huaixing and Li Lingxia and Shi Tian and Zhang Shinan},
title = {Construction of Recombinant Protein of Botulinum Neurotoxin Light Chain and Analysis of Cleavage Effect on VAMP1 Protein in Brown Rat
},
journal = {Animal and Veterinary Sciences},
volume = {12},
number = {6},
pages = {171-176},
doi = {10.11648/j.avs.20241206.14},
url = {https://doi.org/10.11648/j.avs.20241206.14},
eprint = {https://article.sciencepublishinggroup.com/pdf/10.11648.j.avs.20241206.14},
abstract = {In order to explore the feasibility of the new rodenticide of type D Botulinum toxin to prevent and control the population density of pests in farmland from a molecular perspective. This study aimed to analyze the feasibility of D-type botulinum toxin for the control of urban rodents from a molecular perspective, and to detect the cleavage effect of D-type botulinum neurotoxin light chain recombinant protein on the synaptic vesicle membrane protein VAMP-1 of brown house mouse. The genomic DNA of Clostridium botulinum D 8901 strain was used as a template. Based on the botulinum toxin D gene sequence reported in GenBank, specific primers were designed. Plasmids were constructed by gene synthesis and subcloned into the pET28a expression vector. The recombinant plasmid was transformed into BL21 (DE3) Rosetta competent cells, and induced expression by IPTG. The expression products were purified by Ni-NTA affinity chromatography, identified by SDS-PAGE and Western blotting, and the recombinant proteins were analyzed by SDS-PAGE and Western blotting. The cleavage of VAMP1 protein in SD brown house mouse. The pET-28a- BDLc expression plasmid was successfully constructed and transformed into E. coli BL21 (DE3) Rosetta. Western blot confirmed that the recombinant protein pET-28a- BDLc (residues Met 1-Met 94) obtained soluble expression, and obtained 5 mg, Recombinant protein with purity > 90%. It has good biological activity as determined by animal method. The recombinant protein can decompose VAMP1 protein into two fragments. Type D botulinum toxin protein can specifically cleave synaptic vesicle membrane protein (VAMP1) of brown house mouse, and type D botulinum toxin is feasible for urban rodent control.
},
year = {2024}
}
TY - JOUR T1 - Construction of Recombinant Protein of Botulinum Neurotoxin Light Chain and Analysis of Cleavage Effect on VAMP1 Protein in Brown Rat AU - Li Shengqing AU - Han Shengyi AU - Li Shuping AU - Hu Guoyuan AU - Liu Huaixing AU - Li Lingxia AU - Shi Tian AU - Zhang Shinan Y1 - 2024/12/31 PY - 2024 N1 - https://doi.org/10.11648/j.avs.20241206.14 DO - 10.11648/j.avs.20241206.14 T2 - Animal and Veterinary Sciences JF - Animal and Veterinary Sciences JO - Animal and Veterinary Sciences SP - 171 EP - 176 PB - Science Publishing Group SN - 2328-5850 UR - https://doi.org/10.11648/j.avs.20241206.14 AB - In order to explore the feasibility of the new rodenticide of type D Botulinum toxin to prevent and control the population density of pests in farmland from a molecular perspective. This study aimed to analyze the feasibility of D-type botulinum toxin for the control of urban rodents from a molecular perspective, and to detect the cleavage effect of D-type botulinum neurotoxin light chain recombinant protein on the synaptic vesicle membrane protein VAMP-1 of brown house mouse. The genomic DNA of Clostridium botulinum D 8901 strain was used as a template. Based on the botulinum toxin D gene sequence reported in GenBank, specific primers were designed. Plasmids were constructed by gene synthesis and subcloned into the pET28a expression vector. The recombinant plasmid was transformed into BL21 (DE3) Rosetta competent cells, and induced expression by IPTG. The expression products were purified by Ni-NTA affinity chromatography, identified by SDS-PAGE and Western blotting, and the recombinant proteins were analyzed by SDS-PAGE and Western blotting. The cleavage of VAMP1 protein in SD brown house mouse. The pET-28a- BDLc expression plasmid was successfully constructed and transformed into E. coli BL21 (DE3) Rosetta. Western blot confirmed that the recombinant protein pET-28a- BDLc (residues Met 1-Met 94) obtained soluble expression, and obtained 5 mg, Recombinant protein with purity > 90%. It has good biological activity as determined by animal method. The recombinant protein can decompose VAMP1 protein into two fragments. Type D botulinum toxin protein can specifically cleave synaptic vesicle membrane protein (VAMP1) of brown house mouse, and type D botulinum toxin is feasible for urban rodent control. VL - 12 IS - 6 ER -
Qinghai Academy of Animal Science and Veterinary Medicine, Qinghai University, Xining, China; Qinghai Provincial Key Laboratory of Pathogen Diagnosis for Animal Doseases and Green Technical Research for Prevenion and Control, Xining, Chuna
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