Donor Strains
Shuttle vectors maintained in shuttle host donors may be transferred to their recipient hosts by either transformation or by conjugation. Plasmids propagated in the five E. coli hosts listed here can be transferred by either process. With the exception of S17.1, all carry the R-factor R702 (TcR, SmR, SuR, HgR), which is defective in its own transfer but mediates the mobilisation of co-resident plasmid vectors that carry the oriT region of plasmid RK2. In S17.1 (ATCC 47055), the RP4-derived factors needed to mobilise co-resident plasmids have been integrated in the chromosome.
Strain CA434 has a longstanding history of use as a conjugative donor for transfer to Clostridium species [1]. Traditionally, researchers have constructed shuttle vectors in standard cloning hosts, such as Top10, and subsequently transferred the vectors made to CA434 for conjugation into the clostridial recipient. The process may be shortened by using TopSex, representing Top10 that carries R702 (Andrew Dempster, unpublished).
Type IV restriction-modification systems recognise and cut methylated DNA, which can dramatically affect the frequency of DNA transfer. To counter this barrier, two further hosts are available, sExpress [2] which is defective in cytosine methylation (dcm) and strain InterStellar (James Millard, unpublished) which is mutated in both dcm and adenosine methylation (dam). These strains respectively represent the E. coli donor strains NEB Express (NEB) and Stellar (CLONTECH) carrying R702.
Donor strains available at the online shop
References
[1] Purdy D, O’Keeffe TA, Elmore M, Herbert M, McLeod A, Bokori-Brown M, Ostrowski A, and Minton NP. 2002. Conjugative transfer of clostridial shuttle vectors from Escherichia coli to Clostridium difficile through circumvention of the restriction barrier. Mol. Microbiol. 46:439–452.
https://doi.org/10.1046/j.1365-2958.2002.03134.x
[2] Woods C, Humphreys CM, Rodrigues RM, Ingle P, Rowe P, Henstra AM, Köpke M, Simpson SD, Winzer K, Minton NP. A novel conjugal donor strain for improved DNA transfer into Clostridium spp. Anaerobe. 2019; 59:184-191.
https://doi.org/10.1016/j.anaerobe.2019.06.020