Genomic imprinting is an epigenetic phenomenon resulting in parent-of-origin-specific gene expression. Many imprinted genes play an important role in mammalian reproduction by regulating growth and development. Imprinting has traditionally been studied in the placenta and fetus, but imprints can also alter post-natal nutrient transfer through effects in the mammary gland and brain [1]. The insulin gene, INS, is an important growth factor, and in the tammar wallaby, shows tissue-specific paternal expression both pre-natally, in the yolk sac placenta (like eutherians), and post-natally, in the pouch young (PY) liver and adult mammary gland [1, 2, 3]. Previously, a chimeric transcript was identified containing exons from INS and the nearby tyrosine hydroxylase (TH) gene [3]. The start site of the TH-INS transcript showed an approximately equal presence of methylated and unmethylated DNA [3], but it remained to be confirmed whether this was a differentially methylated region (DMR).
To determine the parental origins of methylation at this site, genomic DNA from tammar PY and matched mothers was genotyped. Of 24 PY, two had single nucleotide polymorphisms for which the maternal allele could be determined. Sanger sequencing of cloned bisulfite PCR products was then used to analyse allele-specific methylation in PY liver samples. A total of twenty clones were sequenced which showed an approximately even representation of maternal and paternal alleles. The majority of sequencing reads showed the five CpG sites at this locus to be methylated on both the maternal and paternal copies.
This result suggests that the TH-INS start site is not a DMR. Perhaps parent-of-origin-specific gene expression from this region is regulated by a distal imprint control region [4], or some other aspect of DNA organisation. Addressing the function of post-natal imprinting will be an important step in the field of reproductive biology.