A standardized BioBrick toolbox for the assembly of sequences in mycobacteria

Thaís Larré Oliveira; Anna Stedman; Caroline Rizzi; Jessica Dorneles; Carlos Eduardo Pouey da Cunha; Antonio Sergio Varela Junior; Odir Antônio Dellagostin; Johnjoe McFadden

doi:10.1016/j.tube.2019.07.002

A standardized BioBrick toolbox for the assembly of sequences in mycobacteria

Tuberculosis (Edinb). 2019 Dec:119:101851. doi: 10.1016/j.tube.2019.07.002. Epub 2019 Jul 19.

Authors

Thaís Larré Oliveira¹, Anna Stedman², Caroline Rizzi³, Jessica Dorneles³, Carlos Eduardo Pouey da Cunha⁴, Antonio Sergio Varela Junior⁵, Odir Antônio Dellagostin³, Johnjoe McFadden⁶

Affiliations

¹ Programa de Pós-Graduação em Biotecnologia, Centro de Desenvolvimento Tecnológico, Universidade Federal de Pelotas, Pelotas, RS, Brazil. Electronic address: thais.larre@ufpel.edu.br.
² Department of Microbial and Cellular Sciences, Faculty of Health and Medical Sciences, University of Surrey, Guildford, Surrey, GU2 7XH, United Kingdom; Department of Viral Immunology, The Pirbright Institute, Pirbright, Ash Road, Woking, Surrey, GU24 0NF, United Kingdom.
³ Programa de Pós-Graduação em Biotecnologia, Centro de Desenvolvimento Tecnológico, Universidade Federal de Pelotas, Pelotas, RS, Brazil.
⁴ Programa de Pós-Graduação em Biotecnologia, Centro de Desenvolvimento Tecnológico, Universidade Federal de Pelotas, Pelotas, RS, Brazil; Department of Microbial and Cellular Sciences, Faculty of Health and Medical Sciences, University of Surrey, Guildford, Surrey, GU2 7XH, United Kingdom.
⁵ Instituto de Ciências Biológicas, Universidade Federal de Rio Grande, Rio Grande, RS, Brazil.
⁶ Department of Microbial and Cellular Sciences, Faculty of Health and Medical Sciences, University of Surrey, Guildford, Surrey, GU2 7XH, United Kingdom. Electronic address: j.mcfadden@surrey.ac.uk.

PMID: 31563455
DOI: 10.1016/j.tube.2019.07.002

Abstract

For more than 25 years, recombinant Mycobacterium bovis BCG has been genetically engineered for use as a vehicle for antigen expression and immunomodulation, typically through introducing or deleting a gene from BCG genome. However, BCG transformation efficacy is still unpredictable, and cloning and expression of sequences from mycobacteria is difficult to predict due to the lack of standardization. To overcome such limitations, we have employed the BioBrick format to construct a toolbox of several mycobacterial parts, including coding sequences, reporter genes, selective markers, promoters, and other regulatory sequences. Additionally, we have developed and characterized BioBrick-compatible episomal vectors that are able to replicate in M. bovis BCG to enable expression of heterologous antigens. The availability of a BCG Biobrick toolbox will enable any coding sequence to be optimally expressed in BCG. We believe that this mycobacterial toolbox represents a standardized and useful kit to enhance the efficacy and use of recombinant BCG.

Keywords: BCG; BioBrick; Mycobacterium bovis; Toolbox; Vector; eGFP.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Cell Line
DNA, Bacterial / genetics*
Genes, Bacterial / genetics*
Genes, Reporter
Genetic Vectors
Humans
Mycobacterium bovis / genetics*
Plasmids / genetics*
Promoter Regions, Genetic

Substances

DNA, Bacterial