Embryonic day 13.5 mRNA profiles for wildtype and MIDEAS and DNTTIP1 knockout MEFs

Objective: Perform deep sequencing on mouse embryonic fibroblasts from e13.5 embryos with wildtype, MIDEAS-/- and DNTTIP1-/- genotypes to provide insights into observed embryonic lethality with associated haemolytic anaemia and heart defects. Method: Heterozygous mates of MIDEAS or DNTTIP1 set up and pregnant females culled at e13.5. MEFs were created from embryos and cultured for 4-6 passages before total RNA extraction and deep sequencing (novogene) to obtain mRNA pofiles. FastQ files processed using TRIMGALORE!, STAR, Picards remove duplicates and SAM Tools. R was used for DESeq2 analysis to find differentially regulated genes Results: A large number of both up and down regulated genes were found for both MIDEAS and DNTTIP1 knockout MEFs compared to wildtype. Of these a significant amount were common to both knockouts suggesting a role in the same complex and shared gene targets. A number of these transcripts were associated with angiogenesis and haematopoiesis as well as neual development. Conclusion: Whilst further biological experiments are required to confirm reason for haemolytic anaemia present in knockout embryos there is a strong indication this could be caused by changes in gene regulation cause by the loss of the MIDAC compex MEF mRNA profiles of wildtype and DNTTIP1-/- (duplicate) or wildtype and MIDEAS-/- (triplicate) from e13.5 embryos were generated by deep sequencing using Ilumina novo-seq platform