Molecular & Genomic Surgery Eric M. Wilson 1/5/10.

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Presentation transcript:

Molecular & Genomic Surgery Eric M. Wilson 1/5/10

I am by no means an expert on this subject. If you have questions, I suggest you ask Tom Theruvath or look it up. DISCLAIMER

Molecular Biology

DNA

DNA Replication

Gene Expression

Transcription -DNA strand is used as a template by RNA polymerase for synthesis of a mRNA strand -Occurs in nucleus -3 RNA polymerases - responsible for reading the gene, and building the mRNA strand. It reads only the 3' to 5' strand -Transcription Factors: bind DNA & help transcription of genes -Initiation Factors: bind RNA polymerase & initiate transcription

Translation -mRNA used as a template by ribosomes for synthesis of proteins -Occurs in cytoplasm -Ribosomes have small & large subunits that read mRNA, then bind appropriate tRNAs that have amino acids & eventually make proteins ç class.unl.edu/biochem/gp2/m_biology/animati on/gene/gene_a3.html

Gene Expression

Terminology -Genome: collective term for all genes present in one organism -DNA sequences of 3 billion base-pairs, carried by 23 pairs of chromosomes -25,000 to 30,000 genes, and overall it is 99.9% identical in all people -Genomics: study of genetic material -Proteomics: study of the structure and expression of proteins as well as the interactions among proteins encoded by a human genome

Cell Cycle -G1: most variable, determines cell cycle length -S: most radioresistant -Growth factors affect cell during G1 -Mitosis -Prophase: centromere attachment -Nucleus disappears -Metaphase: csome alignment -Anaphase: csome pulled apart -Telephase: separate nucleus reforms around each set of csomes

Apoptosis

Signal Transduction Pathways

TGF-β

Gene Therapy -Immunotherapy -Herceptin -Chemotherapy -Gleevac -Gene Therapy

Southern Blotting -Technique of transferring DNA fragments from an electrophoresis gel to a membrane support, and the subsequent analysis of the fragments by hybridization with a radioactively labeled probe -Enables reliable and efficient analysis of size-fractionated DNA fragments in an immobilized membrane support -Obtain information about the physical organization of single and multicopy sequences in complex genomes

Northern Blotting -Technique of size fractionation of RNA in a gel and the transferring of an RNA sample to a solid support (membrane) in such a manner that the relative positions of the RNA molecules are maintained -Resulting membrane then is hybridized with a labeled probe complementary to the mRNA of interest -Signals generated from detection of the membrane can be used to determine the size and abundance of the target RNA -Differs from Southern blot: uses mRNA instead of DNA -Only method that provides information regarding mRNA size and has remained a standard method for detection and quantitation of mRNA

Polymerase Chain Reaction

Immunoblotting (Western)

Immunopreciptation

DNA Microarray There are two major application forms for the technology: identification of sequence (gene/gene mutation) and determination of expression level (abundance) of genes.

Cell Culture

Transgenic Mice

Gene Knockout in Mice