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Genetic drift

The impact of health status and genetic quality on phenotype is usually underestimated despite it participates to results reproducibility and reliability.

Genetic drift is the constant tendency of alleles' number to evolve in a population, even in the absence of selective forces.

You can reduce genetic drift's impact by

• Cryopreserving your model: in case of genetic drift or any other problem that might affect your model, you can recover the original strain and pursue your research with a minimal lost of time and money.
• Refreshing your colony every 5-10 generations by backcrossing to the inbred strain.
• Replacing the breeders at most every 2 years. PHENOMIN is importing new breeders from its supplier twice a year.

Reproducibility and safety of your model colonies

As a follow-up to the previous topic along to past few years, most important questions underline the interplay between experimental design and implementation of the 3Rs in practice. Improving experimental quality and optimizing scientific data also goes hand in hand with improving the protocols and techniques we apply in animals or on samples from animal research, which will be processed for data analysis, and especially with reducing the number of animals used. Most we are increasing the robustness of our method more we preserve our mouse lines.   

Reproducibility with 16S metagenomics in microbiome science

16S rRNA gene studies (amplification of hypervariable regions of the 16S region followed by high-throughput sequencing of this PCR product) are widely performed for bacteria identification and quantification. However, many technical challenges are to be considered to ensure good quality and reproducibility of 16S sequencing and analyses. Sample storage and nucleic acid isolation methods influence the composition of the microbiome in the extracted sample. PCR and sequencing errors, filtering algorithms and methods for data analyses impact the veracity and the taxonomic resolution that can be achieved. Finally all these topics consist in the best practices for improving the reproducibility of 16S experiments.

CRISPR/Cas9 genome editing: challenges for research reproducibility and clinical safety

Genome editing tools (especially CRISPR/Cas9) have already revolutionized biomedical research. But what promise do they hold for clinical studies? Several years of using genome editing in basic research have revealed the unpredictable nature and the uncertainty of the mutations that are obtained. While the bearing of off-target effects is widely known (& sometimes overestimated); on-target effects are less recognized and underestimated. Recent work from animal and cell models demonstrates the importance of extensive validation for the risk assessment of genome editing and its impact on cell integrity.

Improve reproducibility and animal welfare with better genotyping practices

Genotyping is an essential step in animal research as it enables the selection of animals that will be bred to amplify a colony (or euthanised to control the space available in the animal house) and of course used in experimental protocols.

Wrongly, genotyping may be considered as an easy step to carry out, but providing accurate and rapid results can be more difficult than one may think. Few data are published on inconclusive genotyping, but it is believed that the classical error rate is close to 10% in mice. Lloyd et al. showed than more 15% of the lines deposited in public repositories such as the MMRRCs and Jackson Laboratory (JAX) do not carry the mutation specified by the depositor. False negative and false positive genotyping can clearly be considered as one of the many factors influencing pre-clinical studies and basic research contributing to the "reproducibility crisis".

As part of the 3Rs improvements, transgenic rodent genotyping is slowly beginning to move from biopsy sampling to different non-invasive methods, such as buccal or anal swabs, faeces or hair follicles. These protocols are rarely applied by scientists for fear of their reliability and non-repeatability and because of the risk of increased contaminations. We have evaluated different non-invasive sampling methods in order to develop a refined and more importantly a robust method that does not increase genotyping errors.

The 3 topics were presented as a learning series features from Charles River short training session during Spring 2021; you can find on-demand the presentations (each no more than 15 minutes) across six different tracks.

• The microbiome: G. Pavlovic had presented "Reproducibility with 16S metagenomics"
• Emerging hot topics: G. Pavlovic had presented "CRISPR/Cas9 genome editing: challenges for research reproducibility and clinical safety" and S. Jacquot had presented  "Improve reproducibility and animal welfare with better genotyping practices"