Abstract
Horizontal gene transfer (HGT) in prokaryotes disseminates genetic information throughout a population and can facilitate adaptation and evolution of the species. Mycobacteria utilize an atypical method of conjugation called distributive conjugal transfer (DCT), which results in mosaic genomes and the potential for accelerated evolution beyond that enabled by the more classical oriT-mediated conjugation. The following is a description of the basic DCT protocol, some possible variations of the assay, and examples of downstream applications to better understand mycobacterial functions.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Frost LS, Leplae R, Summers AO, Toussaint A (2005) Mobile genetic elements: the agents of open source evolution. Nat Rev Microbiol 3(9):722–732. https://doi.org/10.1038/nrmicro1235
Thomas CM, Nielsen KM (2005) Mechanisms of, and barriers to, horizontal gene transfer between bacteria. Nat Rev Microbiol 3(9):711–721
Derbyshire KM, Gray TA (2014) Distributive conjugal transfer: new insights into horizontal gene transfer and genetic exchange in mycobacteria. Microbiol Spectr 2(1):MGM2-0022-2013. https://doi.org/10.1128/microbiolspec.MGM2-0022-2013
de la Cruz F, Frost LS, Meyer RJ, Zechner EL (2010) Conjugative DNA metabolism in gram-negative bacteria. FEMS Microbiol Rev 34(1):18–40. 1111/j.1574–6976.2009.00195.x
Smillie C, Garcillan-Barcia MP, Francia MV, Rocha EPC, de la Cruz F (2010) Mobility of plasmids. Microbiol Mol Biol Rev 74(3):434–452. https://doi.org/10.1128/mmbr.00020-10
Cavalli LL, Heslot H (1949) Recombination in bacteria; outcrossing Escherichia coli K 12. Nature 164(4181):1057
Hayes W (1953) The mechanism of genetic recombination in E. coli. Cold Spring Harb Symp Quant Biol 18:75–93
Wang J, Karnati PK, Takacs CM, Kowalski JC, Derbyshire KM (2005) Chromosomal DNA transfer in Mycobacterium smegmatis is mechanistically different from classical Hfr chromosomal DNA transfer. Mol Microbiol 58(1):280–288
Gray TA, Krywy JA, Harold J, Palumbo MJ, Derbyshire KM (2013) Distributive conjugal transfer in mycobacteria generates progeny with meiotic-like genome-wide mosaicism, allowing mapping of a mating identity locus. PLoS Biol 11(7):e1001602. https://doi.org/10.1371/journal.pbio.1001602
Coros A, Callahan B, Battaglioli E, Derbyshire KM (2008) The specialized secretory apparatus ESX-1 is essential for DNA transfer in mycobacterium smegmatis. Mol Microbiol 69(4):794–808
Flint JL, Kowalski JC, Karnati PK, Derbyshire KM (2004) The RD1 virulence locus of Mycobacterium tuberculosis regulates DNA transfer in Mycobacterium smegmatis. Proc Natl Acad Sci U S A 101(34):12598–12603
Gray TA, Clark RR, Boucher N, Lapierre P, Smith C, Derbyshire KM (2016) Intercellular communication and conjugation are mediated by ESX secretion systems in mycobacteria. Science 354(6310):347–350. https://doi.org/10.1126/science.aag0828
Daffe M, Crick DC, Jackson M (2014) Genetics of capsular polysaccharides and cell envelope (Glyco)lipids. Microbiol Spectr 2(4.):MGM2-0021-2013). https://doi.org/10.1128/microbiolspec.MGM2-0021-2013
Hoffmann C, Leis A, Niederweis M, Plitzko JM, Engelhardt H (2008) Disclosure of the mycobacterial outer membrane: cryo-electron tomography and vitreous sections reveal the lipid bilayer structure. Proc Natl Acad Sci U S A 105(10):3963–3967
Shiloh MU, DiGiuseppe Champion PA (2009) To catch a killer. What can mycobacterial models teach us about Mycobacterium tuberculosis pathogenesis? Curr Opin Microbiol 13:1–7
Boritsch EC, Khanna V, Pawlik A, Honoré N, Navas VH, Ma L, Bouchier C, Seemann T, Supply P, Stinear TP, Brosch R (2016) Key experimental evidence of chromosomal DNA transfer among selected tuberculosis-causing mycobacteria. Proc Natl Acad Sci U S A 113(35):9876–9881. https://doi.org/10.1073/pnas.1604921113
Sapriel G, Konjek J, Orgeur M, Bouri L, Frezal L, Roux AL, Dumas E, Brosch R, Bouchier C, Brisse S, Vandenbogaert M, Thiberge JM, Caro V, Ngeow YF, Tan JL, Herrmann JL, Gaillard JL, Heym B, Wirth T (2016) Genome-wide mosaicism within Mycobacterium abscessus: evolutionary and epidemiological implications. BMC Genomics 17:118. https://doi.org/10.1186/s12864-016-2448-1
Supply P, Marceau M, Mangenot S, Roche D, Rouanet C, Khanna V, Majlessi L, Criscuolo A, Tap J, Pawlik A, Fiette L, Orgeur M, Fabre M, Parmentier C, Frigui W, Simeone R, Boritsch EC, Debrie AS, Willery E, Walker D, Quail MA, Ma L, Bouchier C, Salvignol G, Sayes F, Cascioferro A, Seemann T, Barbe V, Locht C, Gutierrez MC, Leclerc C, Bentley SD, Stinear TP, Brisse S, Medigue C, Parkhill J, Cruveiller S, Brosch R (2013) Genomic analysis of smooth tubercle bacilli provides insights into ancestry and pathoadaptation of Mycobacterium tuberculosis. Nat Genet 45(2):172–179. https://doi.org/10.1038/ng.2517
Parsons LM, Jankowski CS, Derbyshire KM (1998) Conjugal transfer of chromosomal DNA in Mycobacterium smegmatis. Mol Microbiol 28:571–582
Snapper SB, Melton RE, Mustafa S, Kieser T, Jacobs WR Jr (1990) Isolation and characterization of efficient plasmid transformation mutants of mycobacterium smegmatis. Mol Microbiol 4(11):1911–1919
Lee MH, Hatfull GF (1993) Mycobacteriophage L5 integrase-mediated site-specific integration in vitro. J Bacteriol 175(21):6836–6841
Judd J, Boucher N, Van Roey E, Gray TA, Derbyshire KM (2017) Application of distributive conjugal DNA transfer in Mycobacterium smegmatis to establish a genome-wide synthetic genetic array. J Bacteriol. https://doi.org/10.1128/JB.00410-17
Acknowledgments
This work was supported by funding from the NIH (AI042308 and AI07258) and NSF (MCB-1614178).
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2020 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Clark, R.R., Gray, T.A., Derbyshire, K.M. (2020). Quantifying and Characterizing Distributive Conjugal Transfer in Mycobacterium smegmatis. In: de la Cruz, F. (eds) Horizontal Gene Transfer. Methods in Molecular Biology, vol 2075. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9877-7_9
Download citation
DOI: https://doi.org/10.1007/978-1-4939-9877-7_9
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-4939-9876-0
Online ISBN: 978-1-4939-9877-7
eBook Packages: Springer Protocols