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Efficiency comparison of B6(Cg)-Tyr^c-2j /J and C57BL/6NTac embryos as hosts for the generation of knockout mice.

ABSTRACT: Careful selection of the host embryo is critical to the efficient production of knockout (KO) mice when injecting mouse embryonic stem (mES) cells into blastocysts. B6(Cg)-Tyr^c-2j/J (B6 albino) and C57BL/6NTac (B6NTac) strains of mice are widely used to produce host blastocysts for such procedures. Here, we tested these two strains to identify an appropriate match for modified agouti C57BL/6N (JM8A3.N1) mES cells. When comparing blastocyst yield, super-ovulated B6NTac mice produced more injectable blastocysts per female than B6 albino mice (8.2 vs. 5.4). There was no significant difference in birth rate when injected embryos were transferred to the same pseudopregnant recipient strain. However, the live birth rate was significantly higher for B6NTac blastocysts than B6 albino blastocysts (62.7% vs. 50.2%). In addition, the proportion of pups exhibiting high-level and complete chimerism, as identified by coat color, was also significantly higher in the B6NTac strain. There was no obvious difference in the efficiency of germline transmission (GLT) when compared between B6NTac and B6 albino host embryos (61.5% vs. 63.3% for mES clones; 64.5% vs. 67.9% for genes, respectively), thus suggesting that an equivalent GLT rate could be obtained with only a few blastocyst injections for B6NTac embryos. In conclusion, our data indicate that B6NTac blastocysts are a better choice for the microinjection of JM8A3.N1 mES cells than B6 albino blastocysts.

SUBMITTER: Ma Y

PROVIDER: S-EPMC8169498 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The prevalence of seizures in individuals with fragile X syndrome (FXS) is ~25%; however, there are no reports of spontaneous seizures in the Fmr1 knockout mouse model of FXS. Herein, we report that 48% of adult (median age P96), Fmr1 knockout mice from our colony were found expired in their home cages. We observed and recorded adult Fmr1 knockout mice having spontaneous convulsions in their home cages. In addition, we captured by electroencephalography an adult Fmr1 knockout mouse having a spontaneous seizure-during preictal, ictal, and postictal phases-which confirmed the presence of a generalized seizure. We did not observe this phenotype in control conspecifics or in juvenile (age <P35) Fmr1 knockout mice. We hypothesized that chronic, random, noise perturbations during development caused the phenotype. We recorded decibels (dB) in our vivarium. The average was 61 dB, but operating the automatic door to the vivarium caused spikes to 95 dB. We modified the door to eliminate noise spikes, which reduced unexpected deaths to 33% in Fmr1 knockout mice raised from birth in this environment (P = 0.07). As the modifications did not eliminate unexpected deaths, we further hypothesized that building vibrations may also be a contributing factor. After installing anti-vibration pads underneath housing carts, unexpected deaths of Fmr1 knockout mice born and raised in this environment decreased to 29% (P < 0.01 compared to the original environment). We also observed significant sex effects, for example, after interventions to reduce sound and vibration, significantly fewer male, but not female, Fmr1 knockout mice died unexpectedly (P < 0.001). The spontaneous seizure phenotype in our Fmr1 knockout mice could serve as a model of seizures observed in individuals with FXS, potentially offering a new translationally-valid phenotype for FXS research. Finally, these observations, although anomalous, serve as a reminder to consider gene-environment interactions when interpreting data derived from Fmr1 knockout mice.

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Efficiency comparison of B6(Cg)-Tyr^c-2j /J and C57BL/6NTac embryos as hosts for the generation of knockout mice.

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Efficiency comparison of B6(Cg)-Tyrc-2j /J and C57BL/6NTac embryos as hosts for the generation of knockout mice.

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OmicsDI is part of the ELIXIR infrastructure

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Efficiency comparison of B6(Cg)-Tyr^c-2j /J and C57BL/6NTac embryos as hosts for the generation of knockout mice.