Improve Research Reproducibility A Bio-protocol resource
Concise Method
tooltip

Gene complementation

HK Hidenori Konishi
MH Mamoru Hio
MK Masahiro Kobayashi
RT Ryuichi Takase
WH Wataru Hashimoto
request Request a Protocol
ask Ask a question
Favorite

Plasmids for sph#### gene complementation were constructed as follows. DNA fragments containing sph#### were amplified by PCR using the genomic DNA of strain A1 as a template and two synthetic oligonucleotides as primers (sph#### BamHI in-fusion f and sph#### BamHI in-fusion r). Fragments amplified by PCR were cloned into BamHI-digested pKS13 using the In-Fusion HD cloning kit (Clontech), and the resultant plasmids were designated pKS13-sph####. These plasmids were introduced into E. coli strain DH5α cells. The resultant transformed E. coli cells were used to transconjugate strain A1 gene disruptant cells through triparental mating. The gene-complemented cells were screened on the basis of their resistance to antibiotics. Gene complementation was confirmed by DNA sequencing.

Copyright and License information: The Author(s) ©2020
Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

Do you have any questions about this protocol?

Post your question to gather feedback from the community. We will also invite the authors of this article to respond.

0/150

tip Tips for asking effective questions

+ Description

Write a detailed description. Include all information that will help others answer your question including experimental processes, conditions, and relevant images.

post Post a Question
0 Q&A