Mosquito Cell Technologies at the DRSC-BTRR

Cell-based screens with CRISPR-Cas9 technology

Cell-based functional genomics screens using CRISPR-Cas9 technology have opened the doors to new discovery in mammalian cell lines. This approach typically relies on use of viral vectors to deliver the sgRNA library. For cell lines for which equivalent viral transduction systems are not available, or for assays in which delivery via viral vectors is problematic, this had been a barrier.

R. Viswanatha and colleagues in the Perrimon lab pushed past this barrier by developing a new approach to pooled-format CRISPR cell screening to Drosophila cells. The method relies on site-specific recombination rather than viral delivery of sgRNAs. One thing we find exicting about this new technology? It is extensible to additional cell types including cells from other insect species.

Seeking new collaborations for mosquito cell projects

The DRSC-BTRR in the Perrimon lab at Harvard Medical School has among its goals to make it possible for laboratories focused on mosquito research to perform genome-wide, pooled format screens in mosquito cell lines. The most approachable screens are 'selections' that identify genes for which knockout confers resistance to a toxin, drug, or other perturbation that normally kills cells or slows division. However, work in other systems suggests that many other screen assays can be performed using this technology, including flow cytometry-based assays.

We are interested to partner with mosquito biologists and others who can further test the screening approach for use in biomedically relevant assays. For some mosquito species, we are actively developing 'screen-ready' cell lines. For others, we can provide reagents that would make it possible for you to first engineer screen-ready cells, then perform CRISPR-based studies including pooled-format screens.

What's available (as of December 2021)

We have developed and tested a screening-ready cell line for the mosquito species Anopheles Sua5b, as well as focused and genome-wide sgRNA libraries, and demonstrated effectiveness of the screening platform in these cells, using proof-of-concept selection-based assays. In addition, the system is being developed for specific cell lines derived from Aedes aegypti, Aedes albopictus, and Culex quinquefasciatus. As noted above, we can empower others to establish the system in additional cell lines by providing relevant plasmid vectors and protocols.

We have made modified mosquito cell lines compatible with CRISPR screening, as described in Viswanatha, Mameli, et al. (2021) Nature Communications, available at the Drosophila Genomics Resource Center (DGRC) in Bloomington, IN. To request mosquito cell lines, please visit the BDSC website or reach out to their staff. We have made plasmids for mosquito CRISPR available at Addgene. To access the Nat Comm publication, please see below.

In addition, our bioinformatics group, led by Y. Hu, has developed a mutli-functional new online resource, CRISPR Guide Express, relevant to work in mosquitos or mosquito cell lines. One function provides support for ortholog mapping between Drosophila and several mosquito species genes. Another supports identification of pre-computed CRISPR sgRNA designs for multiple mosquito species. The resource also facilitates query of public RNAseq data for mosquito cells, for example to check if a specific gene or set of genes is expressed in a given cell line. Specific Anopheles, Aedes, and Culex species are supported in the sgRNA design resource. A YouTube demo video is available here.

In sum, we have available:

Other species' cells?

Cultured cell lines are available for many insects and other species. If you can imagine the pooled-format CRISPR screening technology we describe being useful in a cell system you are working with, please feel free to get in touch so we can explore together what might be possible. In addition, we have previously established the approach for Drosophila melanogaster cells.

Contact us

  • Questions or comments, including inquiries about new collaborations, can be sent to DRSC-BTRR Director Stephanie Mohr
  • You can also use our bug report form to make a comment or ask a question about an online resource.

Relevant publications