CRISPR modified cell lines
We are using CRISPR gene editing technologies to generate new cell lines as part of the funded project NIH ORIP R24 OD019847 "Next-generation Drosophila cell lines to elucidate the cellular basis of human diseases" (N. Perrimon, PI; A. Simcox, Co-PI).
GFP-tagged knock-in cell lines
As part of the ORIP-funded project, we are making GFP-tagged cell lines, with an emphasis on visualization of various organelles and sub-cellular compartments. The following GFP knock-in cell lines made at the DRSC in collaboration with the Bellne lab are available for distribution by the DGRC in Bloomington, IN.
Note that the parental cell line is positive for an mCherry fusion and Cas9, as the parental cell line is S2R+-MT::Cas9 (DGRC cell catalog #268), which is described in Viswanatha et al. 2018 (PubMed ID 30051818). This parental cell line was itself derived from DRSC cell line S2R+ NPT005 (DGRC cell catalog #229), which is described in Neumuller et al. 2012 (PubMed ID 22174071).
S2R+ with GFP::Cnx99a. Ordering information: DGRC cell catalog ID #273
S2R+ with GFP::Rab11. Ordering information: DGRC cell catalog ID #274
S2R+ with GFP::Polo. Ordering information: DGRC cell catalog ID #275
S2R+ with GFP::Gmap (clone #4). Ordering information: DGRC cell catalog ID #276
S2R+ with GFP::Gmap (clone $7). Ordering information: DGRC cell catalog ID #277
S2R+ with GFP::Fib (clone #11). Ordering information: DGRC cell catalog ID #278
S2R+ with GFP::Fib (clone #12). Ordering information: DGRC cell catalog ID #279
S2R+ with GFP::Golgin. Ordering information: DGRC cell catalog ID #280
S2R+ with GFP::Arl8. Ordering information: DGRC cell catalog ID #291
S2R+ with GFP::Lam. Ordering information: DGRC cell catalog ID #292
S2R+ with GFP::Spin. Ordering information: DGRC cell catalog ID #293
S2R+ with GFP::Sec23. Ordering information: DGRC cell catalog ID #294
These cell lines were made using constructs designed and provided by Kanca and Bellen (Baylor College of Medicine). The cell lines were engineered, isolated, and validated at the DRSC. Validation testing included live-cell imaging, fixed-cell imaging (co-stained with an antibody, when possible), and molecular characterization of the insertion endpoints.
In addition, C-terminal GFP knock-in cell lines were generated as described in a BioRxiv preprint from Bosch et al. (2019) using an 'armless' donor approach. Act5c::GFP, Tub84B::GFP, His2Av::GFP, and Lamin::GFP fusion cell lines were made using this approach and are being shared with the DGRC for distribution to the community.
Knockout cell lines
S2R+-ZnT63C-KO, NHEJ-mediated knockout of ZnT63C. As described in PMID: 29223976. Ordering information: DGRC cell catalog #265.
S2R+-IA2-KO, NHEJ-mediated knockout of ia2. As described in PMID: 29223976. Ordering information: DGRC cell catalog #266.
These cell lines have been sequence verified as containing only knockout alleles by PCR amplification of the target region followed by next-generation sequencing of the PCR amplicon, contig assembly, and comparison with the wild-type reference sequence. We wanted to make sure that the distribution copies of the cell lines are correct. To do this, the DGRC prepared genomic DNA from their distribution copies of the cell lines and shipped that gDNA to the DRSC/TRiP, which we then used as the template for PCR and NGS analysis.
Did you request these cells from the DGRC and use them in a study? If so, please acknowledge both the cell line developers and distribtors by citing NIH Grant 5R24OD019847, which supported production of the resource at DRSC/TRiP, and the Drosophila Genome Resource Center, NIH grant 2P40OD010949, as well as the relevant pulication (manuscript in preparation).