We think you have liked this presentation. If you wish to download it, please recommend it to your friends in any social system. Share buttons are a little bit lower. Thank you!
Presentation is loading. Please wait.
Published byAubrey Carpenter
Modified about 1 year ago
HC70AL Presentation: Gene-Knockout Analysis Arabidopsis Thaliana Genes –AT4g27410 –AT5g07680 Salk Lines –# –# Presented By Tim Schulz
Outline Overall: What will knocking out these genes tell us about how they function? What are the characteristics of the genes and T-DNA inserts? Plant Genotypes? Plant Phenotypes? What Next?
Structure & Location of AT4g27410 UTRExon Promoter UTRIntron Flanking Sequence Exon (bp) FW RV ~ 1.6kb Entire Gene (w/o T-DNA) .5kb (Salk) ~1.4kb Between Primers (w/o T-DNA) 5’ 3’
Structure & Location of AT5g07680 UTRExonIntron UTR Intron Flanking Sequence Exon (bp) FW RV ~750bp (w/o T-DNA) .5kb (Salk) Exon ~ 1.6kb Entire Gene (w/o T-DNA) 5’ 3’
Actual T-DNA Locations AT4G27410 – Different From SALK AT5G07680 – Same as SALK Salk’s Report for AT4G27410: UTRExon Promoter UTRIntron Flanking Sequence Exon (bp) .5kb (Salk)
Actual T-DNA Locations AT4G27410 – Different From SALK AT5G07680 – Same as SALK My Results for AT4G27410 Show: UTRExon Promoter UTRIntron Flanking Sequence Exon (bp) .5kb (Salk) .5kb (Salk)
Actual T-DNA Location: AT4G27410 PCR Odd Results From FW + RV + LBb1: Plants 6-13
Solution: Separate Primers LB+FW, LB+RV, RV+FW
Actual T-DNA Location: AT4G27410 UTRExon Promoter UTRIntron Flanking Sequence Exon (bp) .5kb (Salk) .5kb (Salk) Conclusion After - Band-Size Analysis - Sequencing:
Genotypes: First SALK Line AT4G W/T 2. HET 3. W/T 4. W/T 5. W/T 8. W/T 7. HOMO 6. HOMO 9. HET 10. W/T 11. W/T 12. HET 13. HOMO Plant 1. W/T 2. Heterozygous 3. W/T 4. W/T 5. W/T 6. Homozygous 7. W/T 8. W/T 9. Heterozygous 10. W/T 11. W/T 12. Heterozygous 13. Homozygous
Genotypes: Second SALK Line AT5G07680 Questions: Missing Plants? Smaller Plants?
Phenotypes? Smaller Plants – NAM Related? Missing Plants – Embryo Lethal?
Genotypes: Second SALK Line Plants #
Smaller Plants: Mutants?
Smaller Plants: Mutants? The Test: #10 & #24 Plants # (Hetero DNA for Positive Control) ~750bp T-DNA
Genotypes: Second SALK Line AT5G W/T 2. W/T 3. W/T 4. W/T 5. HET 8. HET 7. (Too Small) 6. W/T 9. HET 10. W/T 11. W/T 12. W/T 13. HET Plant 1. W/T 2. W/T 3. W/T 4. W/T 5. Heterozygous 6. W/T Heterozygous 9. Heterozygous 10. W/T 11. W/T 12. W/T 13. Heterozygous 24. W/T
Phenotypes? Smaller Plants – NAM Related? Missing Plants – Embryo Lethal?
Missing Plants – Embryo Lethal? A nalysis through Microscopy Plant #8 Heterozygous Silique Image 1 All Alive: No Immediate affect 3:1 Alive/Dead Ratio- Seed Development affected Zygotically 1:1 Alive/Dead Ratio- Female Gametophyte affected, post fertilization
Missing Plants – Embryo Lethal? A nalysis through Microscopy Plant #8 Heterozygous Silique Image 2
Missing Plants – Embryo Lethal? A nalysis through Microscopy Plant #8 Heterozygous Silique Image 2 (opened)
Missing Plants – Embryo Lethal? Heterozygous-Embryo Development Early Heart
Missing Plants – Embryo Lethal? Heterozygous-Embryo Development Heart
Missing Plants – Embryo Lethal? Heterozygous-Embryo Development Late Heart
Missing Plants – Embryo Lethal? Heterozygous-Embryo Development Torpedo
Missing Plants – Embryo Lethal? Heterozygous-Embryo Development Walking Stick
Missing Plants – Embryo Lethal? Heterozygous-Embryo Development U-Turn
Where to go from here… Promoter + GFP Multiple Knockouts (determined via microarrays) Other, non-knockout related experiments
Conclusion Primary Goal Know Gene Functionality Primary Method T-DNA Knockout What are the characteristics of the genes and T-DNA inserts? Plant Genotypes? Plant Phenotypes? What Next?
THE FUNCTION OF AT5G04410 (NAC2) AND AT3G10500 (NAC2-like) NAC Family YUYA HIRAI.
Homeobox leucine zipper protein 9 (HAT9) -AT2G Homeodomain(s) -Leucine Zipper Motif -DNA Binding -Dimerization ?? ~ Helix-turn-helix 5’3’ FWRV.
The HAT2 Homeodomain-Like Transcription Factor Family: Genes AT5G47370 and AT4G17460 Bekah Charney HC70AL Spring 2006.
Genes That Direct Transcription Co-activator Proteins : Do they disrupt/alter seed development? Gene 1: At5g09250 Gene 2: At5g09240 Combiz Richard Abdolrahimi.
The MYB and BHLH Transcription Factor Families by Elaine Chiu.
The C3HC4-Type RING Zinc Finger and MYB Transcription Factor Families Matthew Taube June 5, 2008 HC70AL.
Determining Functionality of Arabidopsis Thaliana Genes in Embryo Development Ria Yagnik.
Is My Gene Important for Seed Development in Plants?? Gene: AT3G53370 Jonathan Milgrom Spring 2004.
Zinc Finger CONSTANS- Related and LOB-Domain Containing Genes Nancy Phang June 4, 2004.
Arabidopsis Thaliana A Study of Genes and Embryo Development By Garen Polatoglu.
Genetics & Genotyping By Kristin Gill & Daisy Robinton HC70AL Spring 2009.
What Are the Methods and Approaches Used to Identify and Study Arabidopsis Seed Knock-Out Mutations? Daisy Robinton Matt Emmer Jason Chai.
F-Box Containing Tubby Transcription Factor Family Daisy Robinton Goldberg Lab Spring 2006.
The SET-Domain Containing Protein and MYB-related Families: Genes AT2G05900 & AT1G17460 Kristin Gill HC70AL Spring 2008.
Searching for Genes Important in Seed Development At1g19000 At1g74840 BY: Mike Douglas.
The Trihelix Transcription Factor Family Heather Hernandez.
At5g A mutant phenotype? Emily Eder HC70AL - Spring 2005.
The Role of Genes AT5G48150 and AT2G04890 in Arabidopsis thaliana Seed Development Jennifer Huynh June 8, 2006 Honors Collegium 70AL Professor Bob Goldberg.
What Are the Methods and Approaches Used Study Knock-Out Mutations? Elaine Chiu Nancy Phang June 4, 2009.
What are the Methods and Approaches Used to Identify and Study Arabidopsis Seed Knock- Out Mutations? Eric Newton Garen Polatoglu Rena Schweizer.
HC70AL Final Presentation Chris McQuilkin June 4 th, 2009.
Fig. S1 Mass spectra analysis of XXT4 reaction products demonstrating xylosyltransferase activity towards cellohexaose substrate. Predominant peaks represent.
T-DNA Mutagenesis T-DNA Mutagenesis. Transfer-DNA Mutagenesis: a chemical or physical treatment that creates changes in DNA sequence which can lead to.
HC70AL Spring 2009 An Introduction to Bioinformatics By Brandon Le & Min Chen April 7, 2009.
Mutations in Arabidopsis Exocyst Gene AtSEC8 Jennie Hines Mentor: John Fowler.
A Hypothesis for the function of gene AT4G23180 in A. thaliana By Nicole Foxworth and Deborah Lee (Ether Fowl Ox)
Arabidopsis thealiana AT3G56230 AT4G18650 What are the functions of them? Are they important for seed development? Spring 2004 Mariko Onozawa.
Cloning Promoters Kelli Henry April 27, Mature Green Zygote PreglobularGlobular Transition HeartTorpedoLinear Cotyledon Bending CotyledonLinear.
By Eden Maloney HC70AL Spring 2009 WRKY 59 and Agamous-Like 69 Genes.
Arabidopsis Experiments Developmental Screen and Phase Changes Reverse Genetics PCR Genotyping.
AtPAT10 TIP1 Akr1p1 Akr2p Erf2p Swf1p Pfa5 Pfa3 GODZ HIP14 Pfa4 AtPAT10 TIP1 Akr1p1 Akr2p Erf2p Swf1p Pfa5 Pfa3 GODZ HIP14 Pfa4 AtPAT10 TIP1 Akr1p1 Akr2p.
Suppl Figure S1. mMDH genes in At and t-DNA mutant characterization. A) MDHs in Arabidopsis B) T-DNA insertions available At1g53240 mMDH1 At3g15020 mMDH2.
Sierra Wolfenbarger John Fowler Rex Cole. Like most plants it completes an “alternation of generation” Diploid to Haploid to Diploid Haploid stage is.
Background Introduction By Bensen Fan Kristin Gill Maria Garcia.
Diagnosis with PCR This is a preparation of DNA. We zoomed in a portion of a gene. We know that two primers, Forward and Reverse, will hybridize at specific.
Mapping in Arabidopsis Jeff Long Salk Institute. Cotyledons (seed leaves) Shoot Apical Meristem Hypocotyl (seedling stem) Root Root Apical Meristem.
Cells and cell growth Membranes and cell walls Fertilization and embryogenesis Seed development and dormancy Germination Water transport Water relations.
Introduction of Seed Development and Arabidopsis Embryo Development By Bekah Charney and Thi Nguyen HC70AL Spring 2006.
Heredity, Genetics and DNA Are my favorite things.
Using mutants to clone genes Objectives 1. What is positional cloning? 2.What is insertional tagging? 3.How can one confirm that the gene cloned is the.
Determining the roles of the BTB genes At2g04740, At4g08455, At1g04390, and At2g30600 in Arabidopsis thaliana growth and development. Brandon D. Blaisdell,
+ Intro to Punnett Squares Mrs. MacWilliams Biology.
Seed Development in Arabidopsis thaliana Chris McQuilkin and Eden Maloney.
FERTILIZATION & Intro to Mendelian Genetics. FERTILIZATION
Louis et al. Figure S1 Mean expression intensity Figure S1. MPL1 expression in Arabidopsis organs The Genevestigator tool (http://www.genevestigator.com/)
Gene Regulation: What it is, and how to detect it By Jordan, Jennifer, and Brian.
Arabidopsis Experiments Forward Genetic Screen (Ethylene Insensitive Mutants) Reverse Genetic Screen / PCR Genotyping (H + - ATPase Mutants)
T HE E FFECT OF ATR ON UV- I NDUCED S TEM C ELL D EATH IN A RABIDOPSIS Robert Ursu Dr. John Hays Oregon State University.
Supplementary Figures SHUGOSHINs and PATRONUS protect meiotic centromere cohesion in Arabidopsis thaliana Linda Zamariola 1, Nico De Storme 1, Katrijn.
© 2017 SlidePlayer.com Inc. All rights reserved.