A zebrafish, traditionally used as a vertebrate model organism. Image source: Wiki Commons

Yes, CRISPR has delivered a home run once again, this time in the field of model organisms. Researchers in a number of different groups have used CRISPR in the common zebrafish model of vertebrate development, paving the way for CRISPR-based genetic engineering studies to proceed with gusto.1 The newly published proof of concept for comprehensive genome editing with CRISPR within the zebrafish model means that developmental biologists everywhere will be able to gain unprecedented control over studies linking genetic factors with ontological traits, moving the entire field forward into a new era of research as a result of a completed editing toolkit. The zebrafish model is far more complex than the other simpler eukaryotic model organisms, and so researchers hoping to conduct effective CRISPR studies within zebrafish will need a software package that can handle the added breadth and depth of the new research environment.

Prior to the new research, many CRISPR-mediated developmental studies were hotly anticipated and many studies were pre-planned after the initial theorizing papers were published, meaning that the palpable excitement of ontologists that’s stewed for a few years can finally spring into action.2 3 The researchers used CRISPR’s ability to perform double strand breaks (DSBs) in the genomic DNA of zebrafish, prompting a molecular repair response which allows for the insertion or deletions as desired in keeping with the standard CRISPR methodology. Prior researchers examining CRISPR within the zebrafish model succeeded in insertion of new genes, but didn’t test if insertions and deletions could be performed simultaneously.4 Similarly, other prior research tested the heritability of CRISPR-mediated mutations within the zebrafish model, but hadn’t yet honed allele targeting to the level that subsequent research has made possible.5

Fishing for data

Now that all of the fundamental operations of genomic engineering are working in the zebrafish model, researchers will have to adapt to the new requirements of studying zebrafish development in a polyvariant way. Not every researcher will use every single tool in their toolbox with every experiment, but it’s clear that the door is open to using new combinations of tools that legacy genetic editing technologies precluded.

Prior ontology research within the zebrafish model required the following:

  • Identifying the locus of desired knockout, mutation, or insertion
  • Choosing between knockout, insertion, mutation, or other edit
  • Determining which genome editing technique to use given the locus and the desired operation
  • Making the edit and confirming the edit’s success via reporter genes
  • Tracking embryonic development in light of the experimental edit

The difficulty of legacy genome editing technologies imposed logistical limits on researchers due to their clunkiness and lack of accuracy. Frequently, researchers would need to perform a number of trials of a single experiment just to ensure that the edits had been made properly—a problem that CRISPR blows away completely. A mature CRISPR toolkit is more than a robust single platform for conducting the entire panorama of genetic edits; it’s a streamlined system that works far more reliably and with less up front effort than the systems which it replaces. Multiple CRISPR experiments can be run simultaneously in the same cell due to the high specificity and low organism-level risk that the platform offers.

As if it weren’t enough, CRISPR is accurate enough to edit with high granularity within the same locus.6 This means that entirely new classes of studies within development can be performed, provided that researchers can process the data. For instance, it’s now possible to make mixed developmental zebrafish models which could study the effects of in-organism genetic hybrids—a watershed type of study for the purposes of human disease models.7 Researchers who promise to perform such groundbreaking new work will be the first to have their grant applications approved, so we’re about to witness yet another CRISPR-sparked research goldrush.

In these kinds of newly possible studies, the data requirements are quite a bit more involved than prior developmental studies with zebrafish, now requiring:

  • Identifying the loci of interest
  • Choosing which combinations of multi-target editing operations to perform and in which proportions of genomic material
  • Choosing secondary non functional markers for each type of edit
  • Confirming that DNA replication functions in light of edits
  • Confirming that cells within the blastocyst function in light of edits
  • Confirming that embryonic cells maintain their higher order organization in light of edits
  • Making the edits in experimental zebrafish cells
  • Tracking the proportional edited variants throughout further development and into the mature zebrafish via functional and behavioral traits
  • Tracking heritability of each proportional edits through zebrafish reproduction if necessary
  • Tracking proportional variants in comparison to each other and in comparison to control zebrafish

Navigating an ocean of data

Many developmental biologists aren’t renowned for their love of structured data management or computational processing of data, but they’re going to have to quickly adapt to the present state of affairs caused by CRISPR. Old laboratory management systems may have worked for small scale editing where old data could be ignored or analyzed by an individual, but the scale of experimentation within the zebrafish model is about to get much larger than ontologists are used to. Using legacy systems for laboratory management won’t be able to even plan an experiment utilizing CRISPR within a zebrafish model, nevermind track and share the data, animals, inventory or results. Luckily, there is a powerful software suite which can help with the end-to-end management of massive projects within developmental biology.

Unified Lab Management is the experiment planning, data sharing, and ELN of the CRISPR age. Using Unified Lab Management, you’ll be able to plan and execute massive experiments within animal models with ease. Contact us today to find out how you can use Unified Lab Management to start using the power of CRISPR within your zebrafish models and revolutionize developmental biology.

  1. “Zebrafish Genome Engineering Using the CRISPR-Cas9 System.” December 2016, http://www.cell.com/trends/genetics/abstract/S0168-9525(16)30137-8
  2. “A CRISPR view of development.” 2014, http://genesdev.cshlp.org/content/28/17/1859.full
  3. “Efficient multiplex biallelic zebrafish genome editing using a CRISPR nuclease system.” 2013, http://www.pnas.org/content/110/34/13904.full
  4. “Efficient generation of knock-in transgenic zebrafish carrying reporter/driver genes by CRISPR/Cas9-mediated genome engineering.” 2014, https://www.nature.com/articles/srep06545
  5. “Heritable and Precise Zebrafish Genome Editing Using A CRISPR-Cas9 System.” July 2013, http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0068708
  6. “Genome engineering using the CRISPR-Cas9 system.” October 2013, http://zlab.mit.edu/assets/reprints/Ran_FA_Nat_Protoc_2013.pdf
  7. “CRISPR/Cas9 in zebrafish: an efficient combination for human genetic diseases modeling.” November 2016, https://link.springer.com/article/10.1007/s00439-016-1739-6