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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.
Journal cover image

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