Lecture 11 28.12.2017 – FALL 2017 Modern methods in Molecular Biology Epigenetisc Crispr/CAs9 RNAi

Epigenetics Epigenetic  refers to heritable changes in gene expression (active versus inactive genes), that does not involve changes to the underlying DNA sequence; a change in phenotype without a change in genotype. Or “Epigenetics” refers to covalent modification of DNA, protein, or RNA, resulting in changes to the function and/or regulation of these molecules, without altering their primary DNA sequences.

Micro RNA molecules MicroRNAs (miRNAs) comprise species of short noncoding RNA (18-25 bp) that regulate gene expression post-transcriptionally. Recent studies have demonstrated that epigenetic mechanisms, including DNA methylation and histone modification, not only regulate the expression of protein-encoding genes, but also miRNAs

microRNA processing Picture taken from: http://discovermagazine.com/~/media/import/images/0/2/6/micrornadiag.jpg

Introduction Crispr/Cas9 Clustered regularly-interspaced short palindromic repeats (abbreviated as CRISPR, pronounced crisper) are segments of prokaryotic DNA containing short repetitions of base sequences. CRISPR is being used as a tool that allows scientists to edit genomes with unprecedented precision, efficiency, and flexibility. CRISPR is far better than older techniques for gene splicing and editing

https://www.youtube.com/watch?v=4YK Fw2KZA5o https://www.youtube.com/watch?time_ continue=52&v=0dRT7slyGhs http://www.genetherapynet.com/gene- editing-tools/crispr-cas9.html

Applications Like RNAi, CRISPR interference (CRISPRi) turns off genes in a reversible fashion by targeting, but not cutting a site. The targeted site is methylated so the gene is epigenetically modified. This modification inhibits transcription.. Cas9 was used to carry synthetic transcription factors (protein fragments that turn on genes) that activated specific human genes. CRISPR may be used at the germline level to create animals where the gene is changed everywhere. CRISPR can also be utilized to create human cellular models of disease. For instance, CRISPR was applied to human pluripotent stem cells to introduce targeted mutations in genes relevant to two different kidney diseases, polycystic kidney disease and focal segmental glomerulosclerosis.

RNA Interference (RNAi) supresses the expression of genes, VIRUS defense system Previously known as co-supression analyzes the gene function for drug delivery Involve micro RNA (miRNA) & small interfering RNA (siRNA)

http://www.rxipharma.com/wp-content/uploads/2017/03/sd-rxRNA-Novel-Technology-Platform-1024x607.png

Video link https://www.youtube.com/watch?time_continue =64&v=cK-OGB1_ELE

Summary RNAi≠ CRISPR/Cas9 Silencing and editing DNA Silencing DNA