Experimental Verification Department of Genetic Medicine

Slides:

Advertisements

Similar presentations

PCR Lab Notes. What does PCR Stand For? Polymerase chain reaction.

Advertisements

20,000 GENES IN HUMAN GENOME; WHAT WOULD HAPPEN IF ALL THESE GENES WERE EXPRESSED IN EVERY CELL IN YOUR BODY? WHAT WOULD HAPPEN IF THEY WERE EXPRESSED.

Zinc Finger CONSTANS- Related and LOB-Domain Containing Genes Nancy Phang June 4, 2004.

Alignment of mRNAs to genomic DNA Sequence Martin Berglund Khanh Huy Bui Md. Asaduzzaman Jean-Luc Leblond.

BME 130 – Genomes Lecture 7 Genome Annotation I – Gene finding & function predictions.

BI420 – Course information Web site: Instructor: Gabor Marth Teaching.

Lecture Objectives Define Terms: Transcription, Translation, nucleic acid, amino acid, DNA, RNA, mRNA, cDNA, “ATCG”, Gene, Genomics, Protein, Proteomics,

Syllabus Transcription as the production of mRNA from DNA. The role of RNA polymerase. The splicing of pre-mRNA to form mRNA in eukaryotic cells.

 ribose  Adenine  Uracil  Adenine  Single.

Doug Brutlag 2011 Next Generation Sequencing and Human Genome Databases Doug Brutlag Professor Emeritus of Biochemistry & Medicine Stanford University.

Transcription. the Central Dogma DNA mRNA Protein transcription translation gene expression RPE65 gene RPE65 protein.

Fine Structure and Analysis of Eukaryotic Genes

Analyzing your clone 1) FISH 2) “Restriction mapping” 3) Southern analysis : DNA 4) Northern analysis: RNA tells size tells which tissues or conditions.

Genome Annotation BBSI July 14, 2005 Rita Shiang.

Non-coding RNA Genes the underestimated players in an old game.

Remember the limitations? –You must know the sequence of the primer sites to use PCR –How do you go about sequencing regions of a genome about which you.

Expression of the Genome The transcriptome. Decoding the Genetic Information  Information encoded in nucleotide sequences contained in discrete units.

ModENCODE August 20-21, 2007 Drosophila Transcriptome: Aim 2.2.

Chapter 10 Transcription RNA processing Translation Jones and Bartlett Publishers © 2005.

DNA sequencing. Dideoxy analogs of normal nucleotide triphosphates (ddNTP) cause premature termination of a growing chain of nucleotides. ACAGTCGATTG ACAddG.

LECTURES 3/4. CONSTRUCTING and SCREENING cDNA LIBRARIES to ISOLATE NEW GENES ORIGINAL ARTICLES: CLONING BY COMPLEMENTATION: Lew, D, Dulic, V, and Reed.

DNA TO RNA Transcription is the process of creating a molecule that can carry the genetic blueprint for a particular protein coding gene from the DNA.

1 Transcript modeling Brent lab. 2 Overview Of Entertainment  Gene prediction Jeltje van Baren  Improving gene prediction with tiling arrays Aaron Tenney.

Review of Protein Synthesis. Fig TRANSCRIPTION TRANSLATION DNA mRNA Ribosome Polypeptide (a) Bacterial cell Nuclear envelope TRANSCRIPTION RNA PROCESSING.

The Havana-Gencode annotation GENCODE CONSORTIUM.

Mark D. Adams Dept. of Genetics 9/10/04

Complexities of Gene Expression Cells have regulated, complex systems –Not all genes are expressed in every cell –Many genes are not expressed all of.

Eukaryotic mRNA processing

DNA LIBRARIES Dr. E. What Are DNA Libraries? A DNA library is a collection of DNA fragments that have been cloned into a plasmid and the plasmid is transformed.

While replication, one strand will form a continuous copy while the other form a series of short “Okazaki” fragments Genetic traits can be transferred.

August 20, 2007 BDGP modENCODE Data Production. BDGP Data Production Project Goals 21,000 RACE experiments 6,000 cDNA’s from directed screening and full.

CFE Higher Biology DNA and the Genome Transcription.

Ligate tags SAGE: Procedure Digest with “Tagging enzyme” BsmFI tm Isolate mRNA, RT to cDNA Digest with “Anchoring.

Introduction to molecular biology Data Mining Techniques.

Using DNA Subway in the Classroom Genome Annotation: Red Line.

NAC Family Genes AT1G01720 AT1G77450

Mahmuda Akter, Paige Fairrow-Davis, and Rebecca Seipelt-Thiemann

bacteria and eukaryotes

Answers to Homework Tasks

Information flow from DNA to trait

Lecture 8 A toolbox for mechanistic biologists (continued)

Quiz#3 LC710 9/29/10 name____________

DNA The Molecule of Life.

Supplemental Figure 2. (A) AtplaIVA-1 and AtplaIVA-2 null transcription lines for AtPLAIVA mRNA. RNAs from the relevant wild type Col were isolated.

Supplementary figure 3 a b M 1 2 Probe: Unc5b 7 kb

Expression of the Genome

Chapter 4 “DNA Finger Printing”

Exam #1 W 9/26 at 7-8:30pm in UTC 2.102A Review T 9/25 at 5pm in WRW 102 and in class 9/26.

Expression of the Genome

Gene Editing Design Demo

Genome organization and Bioinformatics

Gene Sizes Vary Strachan p146 DYSTROPHIN.

Project Studying Synechococcus elongatus for biophotovoltaics.

Daily Warm-Up Dec. 11th -What are the three enzymes involved with replication? What is the function of each? Homework: -Read 13.1 Turn in: -Nothing.

closing in on the set of human genes. The ENCODE project.

Figure 3 Transcripts of the splicing mutation (c

Pharmacogenomic variability and anaesthesia

Supplemental Figure 3 A B C T-DNA 1 2 RGLG1 2329bp 3 T-DNA 1 2 RGLG2

Expression of the Genome

Gene Sizes Vary Strachan p146 DYSTROPHIN.

The Structure of the Genome

Hiroaki Matsunami, Linda B Buck Cell

Unit 1: 1.5 Structure of the Genome

Gene Structure.

Identification of Skn-1n, a Splice Variant Induced by High Calcium Concentration and Specifically Expressed in Normal Human Keratinocytes Koji Nakajima,

Expression of multiple forms of MEL1 gene products.

Figure Genetic characterization of the novel GYG1 gene mutation (A) GYG1_cDNA sequence and position of primers used. Genetic characterization of the novel.

Gene Structure.

The Production of Proteins by DNA

Presentation transcript:

Experimental Verification Department of Genetic Medicine Alexandre Reymond Department of Genetic Medicine

Gene predictions outside of VEGA RT-PCR all pairs of exons predicted outside of VEGA objects Goal: Confirm experimentally new gene models Question(s): How good are the predictions outside of the mainstream? Is there any coding gene out there?

Gene predictions outside of VEGA RT-PCR all pairs of exons predicted outside of VEGA objects in 13 of the 44 ENCODE regions Oligos are designed with Primer 3 and default parameters Individually on 24 human polyA RNA

Gene predictions outside of VEGA 1255 pairs of exons predicted by nine different programs outside of VEGA objects (13 ENCODE regions)

Gene predictions outside of VEGA Tested 1215 pairs for a total of 29160 reactions 2.6% of success rate (32/1215) 1.9% with perfectly predicted exon boundaries (23/1215) 0.7% with displaced exon boundaries (9/1215)

Gene predictions outside of VEGA RT-PCR all pairs of exons predicted outside of VEGA objects Goal: Confirm experimentally new gene models Question(s): How good are the predictions outside of the mainstream? Is there any coding gene out there?

RT-PCR VEGA objects RT-PCR of all the VEGA objects belonging to the « novel transcripts », the « putative transcripts» and the « non-canonical splicing » class. Goal: Confirm experimentally new genes Question: Is the VEGA annotation team doing a good job?

5’ RACE 5’ RACE of all the VEGA objects belonging to the « known » and the « novel protein» class Goal: Find new 5’ end exons and/or new 5’ UTRs due to alternative splicing Question: Should a 5’RACE effort be pursued for the entire genome (ENCODE consolidation phase)?

5’ RACE DNA mRNA Primers are designed with “Primer 3” just 3’ of the ATG in an exon common to most (all) described transcripts of this gene Parameters (25 nts, 68°<Tm<72°, best 70°, 50-70% GC) cDNA is prepared with “BD SMART RACE” kit Touchdown 72°, 70° and 68°

5’ RACE results 5’RACE in 12 tissues on 426 genes Pick tissue with a single large bands 52% success rate (220/426) Pick a second tissue with a single large band Add 6% and get to 58% rate success (245/426) Clone and sequence band from tissues with multiple hits Add 5-15%...