Generation of a novel Stra8-driven Cre recombinase strain for use in pre-meiotic germ cells in mice

Abstract

The development of oocytes occurs over a broad time frame, starting at the earliest stages of embryogenesis and continuing into adulthood. Conditional knockout technologies such as the Cre/loxP recombination system are useful for analyzing oocyte development at specific stages, but not every time frame has appropriate Cre drivers, for instance, during oocyte meiotic initiation through early prophase I in the embryo. Here, we generated a novel knockin mouse line that produces a bicistronic transcript from the endogenous Stra8 locus that includes a 'self-cleaving' 2A peptide upstream of cre. This allows for high efficiency cleavage and production of both proteins individually and results in expression of cre in both male and female gonads at the biologically relevant stage. Fluorescent reporter analysis confirms that this line recapitulates endogenous Stra8 expression in both sexes and does not affect fertility of heterozygous nor homozygous mice. This line, named Stra8P2Acre, adds to the repertoire of germ-cell specific cre driver lines and, importantly, allows for deletion of target genes during key embryonic oocyte developmental stages, including early events in meiosis.

Unlocking Genetic Mysteries during the Epic Sperm Journey toward Fertilization: Further Expanding Cre Mouse Lines

“…Avery et al knocked in engineered P2A-Cre via the CRISPR/Cas9 system and recombinase activity without disturbing Stra8 expression or mouse fertility. It is worth noting that the Stra8-P2ACre line also exhibited Cre activity in female mouse germ cells at E12.5-E16.5 [43]. Xue et al generated germline-specific Nat10 cKO females using Stra8-GFPCre or Zp3-Cre KI and revealed Nat10-indispensable oocyte meiosis progression, growth, and maturation [44].…”

“…Moreover, random insertions and uncontrolled gene copies caused by transgenic steps in conventional recombination are avoided in the CRISPR/Cas9 system [197]. In this review, we summarized several genedisrupted mouse lines edited by the CRISPR/Cas9 approach, including Ddx4 iCre [15], Ddx4 em1(CreERT2)Utr [20], Stra8 P2A-Cre [43], Acrv1-iCre [106], Lcn8-Cre and Lcn9-Cre [190,191], which showed higher specificity, efficiency, and the lower occurrence of global KO events. Other recommendations are as follows: 1.…”