Standard Laboratory Strains

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Background and Goal

Dear Colleagues,

The stock center has of course followed the recent discussion about strains with great interest. A lot of good arguments were made.

We want you to know that we are able to help out with whatever you decide. One of the functions of the stock center always has been to provide standardized strains.

We have already had a couple of requests from new laboratories about what strain we would recommend as the lab strain. We are not prepared to give an unequivocal answer, but we would recommend that it be a strain with less (least) amount of duplication. This also is suggested in a paper, soon to be published, by Bloomfield et al (PMID: 18430225). This paper is a study with data concerning duplications and deletions in various laboratory stocks.

Another consideration should be the problems to be studied and with what strains one wants to compare results. For example, a large number of cytokinesis-related studies have been performed with the standardized strain AX2-214 and it might make sense to use this strain for related problems. However, a lot chemotaxis studies have been done with DH1 or AX3 from the Devreotes lab.

One final consideration should be the fact that it is the strain AX4 that has been sequenced.

But standardized strains only work if the labs that use them follow proper culture and maintenance procedures. The most important aspect of this is to store a (large) number of aliquots of axenically grown cells, either at -80C or in liquid nitrogen, as soon as you receive the strain. Every month an aliquot should be thawed to start a new culture.

Another option is to store a few vials of spores at -80C or even at -20C. Every month one can scrape a few spores onto a bacterial plate, and axenic cultures can be started by transfering growing cells from the bacterial plate into HL5 with streptomycin. Protocols are on the dictyBase website.

If you are in the habit of sub-culturing the same stock for months on end, you are definitely not working with the same strain anymore with which you started.

Best regards, Jakob Franke and dictyBase




Genetics of laboratory strains

In recent years some progress has been made in characterising the genetics of the NC4 lineage, which includes most of the standard lab strains. Some unpublished data were presented and discussed at the Annual Dicty Conference held at Potsdam in August 2014, and I will briefly outline some key points which might be of interest to members of the community who were not in attendance (Gareth Bloomfield, 22nd August 2014).

AX2 and AX3 (along with the several AX3-derived strains such as AX4/KAX3, DH1, and JH10) derive from DdB, which was itself selected from NC4 in the Sussman laboratory in the 1960s as a variant that develops synchronously and forms well-defined plaques when grown in association with bacteria on SM agar plates (Jakob Franke, 1999). The genotype of DdB and its descendents could be distinguished from NC4 and other wildtype strains decreases in copy number at a small number of loci (Bloomfield et al., 2008). These copy-number changes represent deletions of genes that are single-copy in NC4, and which are currently under investigation (unpublished data).

Recent work has highlighted problems asociated with the use of axenic mutants when studying chemotaxis in vegetative cells (Veltman et al., 2013). Characterisation of the axeB gene indicates that this is caused at least in part by misregulation of Ras signalling (manuscript in preparation). This, as well as the presence of other mutations shared in AX2 and AX3 lines that are not yet characterised, has prompted renewed efforts to promote the use of wildtype strains in applications for which axenic growth is unnecessary.

Many wildtype D. discoideum strains are available, and any can in principle be used as a general laboratory strain, as earlier discussed in the Strain History email exchange (note especially the contribution from Keith Williams). NC4 lines have advantages, notably the growth phenotypes of DdB; the availability of the well annotated genome of AX4, which differs little from its ultimate parent; and its status as the formal type of the species. However the fact that it was the first isolate is also its major disadvantage: when Raper first cultured the strain he cannot have expected that so many experimentalists would still want to use it more than eighty years later. NC4 was isolated in 1933 (Raper, 1935), and Raper later recorded that it was 'cultivated in association with Vibrio alkaligenes for many months before being shifted to Escherichia coli...' (Raper, 1984). Experiments testing the survival of spores after lyophilization were recorded as having begun in 1941, eight years after the isolation of NC4 and four years after that of another early isolate, V12 (Raper, 1951). It seems reasonable to assume that some genetic changes occurred in the period before long-term storage was initiated. The large segmental duplications identified in different NC4 lines (but not in DdB) by comparative genomic hybridisation (Bloomfield et al., 2008) might reflect chromsome instablility caused by an underlying defect in NC4, or might just be an inevitable consequence of clonal propagation. Whatever the cause, the presence of such duplications is unhelpful in a strain to be used in genetic analysis.

DdB remains a possible choice as a strain lacking duplications, along with other more recent isolates. The Raper laboratory gathered many further isolates (eg Erdos et al., 1973), as did Francis (Francis and Eisenberg 1993) and more recently the laboratory of Queller and Strassman (Douglas et al., 2012).




To collaborate

If you are interested in collaborating on this (ie, use one of the standard strains and make new strains using that stock), please enter your name and the strains you want to work with; or email Pascale (pgaudet@northwestern.edu) or dictyBase (dicty@northwestern.edu) for your name to be added here.

  • Pierre Cosson: AX2
  • Thierry Soldati : AX2
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