# Modular gene tagging in C. elegans

## TL;DR

This paper introduces PhIT, a new method for tagging genes in C. elegans using recombinases, allowing for modular and error-free protein tagging.

## Contribution

The novel contribution is PhIT, a recombinase-based system for modular and germline-competent gene tagging in C. elegans.

## Key findings

- Eight recombinases were tested for germline function in C. elegans.
- PhIT enables insertion of various tags, including fluorescent proteins and degron tags, via genetic crosses.
- The method uses a 39bp PhiC31 attB landing pad inserted by CRISPR as a reusable resource.

## Abstract

Tagging a gene endogenously can identify when the gene is expressed and where the protein is localized. CRISPR is the primary tool for generating tags of endogenous genes, but it is error-prone and requires unique reagents for each gene and tag. Recombinases can insert DNA in an error-free and modular manner. Here, we tested eight recombinases for germline function in the nematode C. elegans, and introduce PhIT, a recombinase-based method for protein tagging. First, a short 39bp PhiC31 attB landing pad is inserted into the locus by CRISPR. This strain is a resource which can be used to insert a variety of modular tags. Second, tags are inserted by the integrase PhiC31, and in tandem, extraneous backbone sequences are removed by a tyrosine recombinase. Current modular tags include seven different fluorescent proteins, FLP-regulated cell-specific expression constructs, and degron tags. Importantly, tags can be inserted by genetic crosses instead of by microinjection.