CSB349H1 Lecture Notes - Lecture 7: Yeast Artificial Chromosome, Olfactory Receptor Neuron, Olfactory Receptor

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Lecture 7(b): Methylation Dynamics & Monoallelic Expression
Question After formation of the blastocyst, we have a re-methylation event; how can some genes
escape this methylation (i.e. housekeeping genes)
DNA Methyltransferases (DNMT):
DNMT3 has enzymatic activity that allows them to methylate DNA
DNMTLdoes not have enzymatic activity but is capable of binding to DNA and recruit
DNMTs to DNA and have them methylated
1. Protection of CpG islands of housekeeping genes from de novo
methylation:
o Histone modification is often found on the promotor
of housekeeping gens: H3K4me
§ Modification functions to inhibit the contact
between DNMT3 and nucleosomes therefore, not allowing methylation
Monoallelic Gene Expression:
Certain genes in the mammalian genome are expressed from only one of the 2 alleles
o Monoallelic expression can be imprinted (paternal/maternal) or random
§ Random => (1) X-linked Genes | (2) Autosomal genes
1. Randomly Expressed Autosomal Genes:
o Certain cells express either maternal or parental allele however, some cells can be
found expressing both (parental + maternal) in the same cell
§ Once a pattern of expression is established it is maintained in all subsequent
cellular clones
ð SNP Analysis revealed that 5-10% of autosomal genes are expressed monoallelically
Detecting Monoallelic Expression:
1. Single Nucleotide Polymorphisms (SNPs)/ RT-PCR:
o Each gene consists of a maternal and paternal allele copy these two copies are often
not identical; slight differences can be found = SNPs
Experiment:
Mouse strains were used to generate an inbred line; homozygous (both copies are identical)
o (M) Parent 1: 4th position is a G | (P) Parent 2: 4th position is an A
§ Crossing the two mouse will generate an offspring with both copies (A&G)
ð Completing an RT-PCR (detecting RNA levels) from offspring’s tissue will display two peaks:
o Two distinct clonal cell lines can be generated: one will
express maternal (G) and other cell line will express
paternal (A)
ð Single Nucleotide Polymorphisms (SNPs) can be used to
distinguish between maternal and paternal alleles
o SNP analysis display different band size
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Document Summary

Question after formation of the blastocyst, we have a re-methylation event; how can some genes escape this methylation (i. e. housekeeping genes) Dna methyltransferases (dnmt): dnmt3 has enzymatic activity that allows them to methylate dna, dnmtl does not have enzymatic activity but is capable of binding to dna and recruit. Dnmts to dna and have them methylated: protection of cpg islands of housekeeping genes from de novo methylation, histone modification is often found on the promotor of housekeeping gens: h3k4me. Modification functions to inhibit the contact between dnmt3 and nucleosomes therefore, not allowing methylation. Monoallelic gene expression: certain genes in the mammalian genome are expressed from only one of the 2 alleles, monoallelic expression can be imprinted (paternal/maternal) or random. Random => (1) x-linked genes | (2) autosomal genes: randomly expressed autosomal genes, certain cells express either maternal or parental allele however, some cells can be found expressing both (parental + maternal) in the same cell.

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