Single cell microarray analysis of mouse primordial germ cells with Prdm14 mutation.. Single cell microarray analysis of mouse primordial germ cells with Prdm14 mutation.

Prdm14 is a critical gene for specifying mouse primordial germ cells (PGCs). The changes in expression in mouse PGCs caused by mutations of the Prdm14 gene were investigated at the single-cell level using microarray analysis. Overall design: C57BL/6 mice with the Prdm14 (+/-) genotype (Yamaji et al., Nature Genetics, 2008) were crossed. Pregnant mice were sacrificed at embryonic day (E) 7.25 - 7.5 and embryos at the early-to-mid allantois bud stage were isolated. The base of the allantois bud, where primordial germ cells (PGCs) were clustered, was dissected using a glass knife and cells were dissociated with trypsin as described (Kurimoto et al., Genes and Development, 2008). Single cells were randomly isolated manually using a glass capillary as described (Kurimoto et al., Nature Protocols, 2007) and directly subjected to cDNA amplification (Kurimoto et al., Nucleic Acids Research, 2006). cDNAs of PGCs were identified based on Prdm1 expression using targeted PCR as described (Kurimoto et al., Genes and Development, 2008). Genotypes of Prdm14 were identified by PCR as described (Yamaji et al., Nature Genetics, 2008). cDNAs of PGCs with Prdm14 (+/+), (+/-), and (-/-) genotypes were then subjected to microarray analysis using the Affymetrix GeneChip Mouse Genome 430 2.0 Array. Microarray data was processed using dChip (Ver.1.3) with the PM/MM difference model. Expression level of Dppa3 (1424295_at) were investigated.