Resources

Neural marker antibodies

CRISPR/Cas9 Protocol

You can find our current CRISPR protocol here. This protocol was developed with lots of support from the Khokha lab and the NXR. Please cite this, this and this.

We get our Cas9-NLS protein from MacroLabs at UC Berkeley.

Genotyping Protocol

We use a CRISPR genotyping protocol to estimate sgRNA efficiencies. You can find our current genotyping protocol here. This is a genomic DNA extraction followed by a PCR that we send for Sanger sequencing and then upload for sequence deconvolution. 

Whole-Mount Immunostaining Protocol

This is great for staining stage 44+ tadpoles (X. tropicalis). We use a high-throughput staining setup (see baskets, racks below), but our protocol also works great in Eppendorf tubes with manual washes. Please cite this and also this! For staining of earlier stage embryos/tadpoles or for X. laevis, you need to add some permeabilization steps— contact Helen for more info (helen.willsey@ucsf.edu).

3D printer files for embryo/tadpole stamps

These are great for positioning animals. Just stamp them into liquified agarose and let it set.

Tadpole stamp (X. tropicalis, stages 44-47)

Embryos stamp (X. tropicalis, stages 1-20)

 Tadpole stamp (X. laevis, stages 44-47)

These were originally conceived of by Richard Harland and Marta Truchado-Garcia, though these files are our creations. Please cite this and this.

3D printer files for ISH/IF basket racks

These are great for high-throughput staining, and don't require a table saw!

Small rack (fits 24 baskets), for glass dish use #900170 from Wheaton Inc.

Medium rack (fits 30 baskets), for glass dish use #900203 from Wheaton Inc.

Large rack (fits 60 baskets), for glass dish use #49WF37 from Grainger

Making Baskets Protocol, we use 200um mesh for IF (Spectra Mesh #146487) and 100um mesh for ISH (Spectra Mesh #146488)

This concept was originally developed by Richard Harland and Karen Liu, though the 3D printer files are our creations. Please cite the basket protocol and our protocol.

RNA-Seq from dissected X. tropicalis brains over development

Click here to explore our X. tropicalis brain time-course RNA-Seq dataset. Bulk sequencing from dissected whole brains from stages 40-47. Please cite it.

Clonotator

Clonotator is a web-based platform made by the Jeremy Willsey lab that integrates several bioinformatics tools to facilitate efficient screening and annotation of cDNA construct sequences for use in molecular biological experiments. Clonotator automatically translates the nucleotide sequence of the construct into an amino acid sequence, aligns the predicted sequence to a reference database of protein sequences and identifies the best protein and isoform match, annotates any variants present in the construct, and incorporates disease-associated mutations and transcriptomic data to help the user assess suitability for a particular biological context (e.g., isoform affected by disease-associated variant, isoform expressed in tissue of interest).