Biochemical Characterization and Functional Analysis of Glucose Regulated Protein 78 from the Silkworm Bombyx mori
Abstract
:1. Introduction
2. Results
2.1. Cloning and Characterization Analysis of BmGRP78 from Bombyx mori
2.2. Expression and Production of Recombinant BmGRP78 (rBmGRP78) and Preparation of Antiserum
2.3. Analysis Expression Patterns of BmGRP78 in Developmental Stages and Larval Tissues
2.4. The ATPase Activity of rBmGRP78
2.5. rBmGRP78 Inhibits Thermal Aggregation of Citrate Synthase (CS) and Malate Dehydrogenase (MDH)
2.6. BmGRP78 Expression in Response to Abiotic and Biotic Stresses in BmN Cells
2.7. Subcellular Localization of BmGRP78 under Different Stress-Induced Conditions in BmN Cells
3. Discussion
4. Materials and Methods
4.1. Biological Materials
4.2. Cloning and Sequence Analysis of BmGRP78
4.3. Bio-Informatic Analysis
4.4. Prokaryotic Expression, Purification, and Antiserum Preparation of Recombinant BmGRP78 (rBmGRP78)
4.5. RNA Extraction and Quantitative Real-Time PCR (qRT-PCR)
4.6. SDS-PAGE and Western Blotting
4.7. The ATPase Activity Assay
4.8. Thermal Aggregation Assays
4.9. Abiotic and Biotic Treatment
4.10. Subcellular Localization by Confocal Microscopy
4.11. Statistical Analysis
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Gene | Primer Sequence (5′-3′) | Application | Length of Product (bp) | |
---|---|---|---|---|
BmGRP78 | Forward | ATGGTCAAGATGCG | ||
Reverse | CAACTCGTCCTTGAAG | |||
Forward | CCCAAGCTTGCCACCACCACCACCACCACATGGTCAAGATGCG | PCR | 2012 | |
Reverse | CCGCTCGAGCAACTCGTCCTTGAAG | |||
Forward | AAGGACATCGGCACAGTAATCG | qRT-PCR | 141 | |
Reverse | ATCTTGAGTGAAGGCCACGTAT | |||
RP49 | Forward | TGCTCCCAAATGGATTCCGTAAG | qRT-PCR | 131 |
Reverse | CACGATCAGCTTCCGCTTCTTC |
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Xiao, Y.; Ren, L.; Wang, Y.; Wen, H.; Ji, Y.; Li, C.; Yi, Y.; Jiang, C.; Sheng, Q.; Nie, Z.; et al. Biochemical Characterization and Functional Analysis of Glucose Regulated Protein 78 from the Silkworm Bombyx mori. Int. J. Mol. Sci. 2023, 24, 3964. https://doi.org/10.3390/ijms24043964
Xiao Y, Ren L, Wang Y, Wen H, Ji Y, Li C, Yi Y, Jiang C, Sheng Q, Nie Z, et al. Biochemical Characterization and Functional Analysis of Glucose Regulated Protein 78 from the Silkworm Bombyx mori. International Journal of Molecular Sciences. 2023; 24(4):3964. https://doi.org/10.3390/ijms24043964
Chicago/Turabian StyleXiao, Yao, Lujie Ren, Yanan Wang, Huanhuan Wen, Yongqiang Ji, Chenshou Li, Yangqing Yi, Caiying Jiang, Qing Sheng, Zuoming Nie, and et al. 2023. "Biochemical Characterization and Functional Analysis of Glucose Regulated Protein 78 from the Silkworm Bombyx mori" International Journal of Molecular Sciences 24, no. 4: 3964. https://doi.org/10.3390/ijms24043964