I plan to buy commercial competent cells, but I am concerned about running out. If I obtain the appropriate strain, can I culture and produce more competent cells from the commercial ones?
That's what most people do. The competent cells are not modified when they become "competent" so you can grow them normally, make glycerol stocks and prepare your cells. You just need a good protocol for competent cell preparation. One advice: plate the cells on agar and pick multiple colonies that look good (grow normal, no strange color etc.). Starting from a single clone can sometimes lead to weird results because you could pick a "weird" clone. Inoculating from glycerol stock directly is possible but has some risk of contamination if you do it repeatedly from the same stock.
Yes you can. There are many protocols out there to make electrocompetent cells.
I am not an expert in this field, but I am very interested and have researched to find an answer. I received some assistance from tlooto.com for this response. Could you please review the response below to see if it is correct?
Yes, you can culture commercially purchased BL21 competent cells and use a chemical method to produce more competent cells. After obtaining the initial batch, you can grow the cells in LB medium, collect them at the exponential growth phase, and induce competence using calcium chloride treatment or other chemical methods. This allows you to produce additional competent cells for future experiments. However, be mindful of potential variability in transformation efficiency and ensure sterile techniques to maintain the strain's integrity [1][2][3].
Reference
[1] Kim, S., Jeong, H., Kim, E., Kim, J. F., Lee, S., & Yoon, S. (2017). Genomic and transcriptomic landscape of Escherichia coli BL21(DE3). Nucleic Acids Research, 45, 5285 - 5293.
[2] Sun, X., Zhang, Z., Wang, L., Wang, J., Liang, Y., Yang, H., Tao, R., Jiang, Y., Yang, J., & Yang, S. (2020). Downregulation of T7 RNA polymerase transcription enhances pET‐based recombinant protein production in Escherichia coli BL21 (DE3) by suppressing autolysis. Biotechnology and Bioengineering, 118, 153 - 163.
[3] Corless, E., Mettert, E. L., Kiley, P., & Antony, E. (2019). Elevated Expression of a Functional Suf Pathway in Escherichia coli BL21(DE3) Enhances Recombinant Production of an Iron-Sulfur Cluster-Containing Protein. Journal of Bacteriology, 202.
yep, that sounds perfect. Check the protocol below:
https://health.uconn.edu/li-lab/wp-content/uploads/sites/129/2017/06/electrocompetent_cells.pdf
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