NTR366L_spring2011_week8_lecture-RTPCR

NTR366L_spring2011_week8_lecture-RTPCR - Todays topics Gene...

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1 Today’s topics Gene Expression Reverse Transcription PCR (RT-PCR) Microarray Models Animals in research Transgenic mice
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2 Central Dogma DNA (Genetic Info) mRNA (Template) Protein (Product) transcription translation
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Definition of gene expression 3 Conversion of the information encoded in a gene first into messenger RNA (mRNA) and then to a protein. Note: Laboratory Detection of gene expression: detect mRNA in a given sample
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What affects Gene Expression Level (mRNA level)? When: embryonic stages/adult/disease progression Where: different tissue Regulators: Transcriptional factors Signaling pathway Methylation (epigenetics) Exogenous agents etc. 4
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mRNA to cDNA cDNA: complementary DNA Transcription: DNA->mRNA Reverse transcription: mRNA->DNA Reverse transcriptase: Biological tool from retrovirus 5
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6 Detection of Gene expression (mRNA) Northern blots ( RNA ) Dot blots ( RNA ) Ribonuclease protection assay (RPA) ( RNA ) In situ hybridization ( RNA ) Reverse transcriptase PCR (RT-PCR) Reverse transcription PCR ( cDNA ) Gene expression microarrays cDNA or oligo probes ( cDNA )
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Note: RNA is extremely susceptible to degradation by nucleases (Rnase) and must be handled with great care! Be Extremely Clean! 7
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8 THIS WEEK’S LAB: One-step Reverse transcriptase PCR ( RT -PCR) Two-steps: Step 1: RT Synthesize complementary DNA (cDNA) from mRNAs using reverse transcriptase Step 2: PCR using Taq DNA polymerase “One-step” RT-PCR Reverse transcriptase + Taq
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Personalized Nutrition Nutrigenetics Effect of gene variants on functional ability Digestion, absorption, use of food Normal functions as well as disorders (PKU, etc.) Nutrigenomics Effect of food on gene expression and function Risk of/susceptibility to disease Diabetes, heart disease, etc. Multiple genes affected
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This note was uploaded on 05/10/2011 for the course NTR 366L taught by Professor Zhu during the Spring '11 term at University of Texas.

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NTR366L_spring2011_week8_lecture-RTPCR - Todays topics Gene...

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