MegaGate: A toxin-less gateway molecular cloning tool.
Publication
, Journal Article
Kramme, C; Plesa, AM; Wang, HH; Wolf, B; Smela, MP; Guo, X; Kohman, RE; Chatterjee, P; Church, GM
Published in: STAR protocols
December 2021
Gateway cloning employs the use of the ccdb toxin and has low colony numbers, making it difficult to apply at scale to clone libraries of cDNA vectors. In this protocol, we describe MegaGate, a toxin-less Gateway technology capable of robust cDNA library cloning that is efficient, cheap, and scalable. MegaGate eliminates the ccdb toxin used in Gateway recombinase cloning and instead utilizes meganuclease-mediated digestion to eliminate background vectors during cloning and is 99.8% efficient with high colony numbers. For complete details on the use and execution of this protocol, please refer to Kramme et al. (2021).
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Published In
STAR protocols
DOI
EISSN
2666-1667
ISSN
2666-1667
Publication Date
December 2021
Volume
2
Issue
4
Start / End Page
100907
Related Subject Headings
- Recombinant Fusion Proteins
- Polymerase Chain Reaction
- Plasmids
- Gene Library
- Escherichia coli
- DNA, Complementary
- Cloning, Molecular
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Kramme, C., Plesa, A. M., Wang, H. H., Wolf, B., Smela, M. P., Guo, X., … Church, G. M. (2021). MegaGate: A toxin-less gateway molecular cloning tool. STAR Protocols, 2(4), 100907. https://doi.org/10.1016/j.xpro.2021.100907
Kramme, Christian, Alexandru M. Plesa, Helen H. Wang, Bennett Wolf, Merrick Pierson Smela, Xiaoge Guo, Richie E. Kohman, Pranam Chatterjee, and George M. Church. “MegaGate: A toxin-less gateway molecular cloning tool.” STAR Protocols 2, no. 4 (December 2021): 100907. https://doi.org/10.1016/j.xpro.2021.100907.
Kramme C, Plesa AM, Wang HH, Wolf B, Smela MP, Guo X, et al. MegaGate: A toxin-less gateway molecular cloning tool. STAR protocols. 2021 Dec;2(4):100907.
Kramme, Christian, et al. “MegaGate: A toxin-less gateway molecular cloning tool.” STAR Protocols, vol. 2, no. 4, Dec. 2021, p. 100907. Epmc, doi:10.1016/j.xpro.2021.100907.
Kramme C, Plesa AM, Wang HH, Wolf B, Smela MP, Guo X, Kohman RE, Chatterjee P, Church GM. MegaGate: A toxin-less gateway molecular cloning tool. STAR protocols. 2021 Dec;2(4):100907.
Published In
STAR protocols
DOI
EISSN
2666-1667
ISSN
2666-1667
Publication Date
December 2021
Volume
2
Issue
4
Start / End Page
100907
Related Subject Headings
- Recombinant Fusion Proteins
- Polymerase Chain Reaction
- Plasmids
- Gene Library
- Escherichia coli
- DNA, Complementary
- Cloning, Molecular