The Goal Put together the parts we have now (and some parts we’re still missing) to make: LacI, TetR, and cI Lambda coding regions w/ hi/low constitutive.

Slides:

Advertisements

Similar presentations

Design and Testing of a Bacteriological Timer Device Herman Armstrong Morgan Schiermeier Rachel Klapper Jackie Schneider.

Advertisements

Genetic Jigsaw Class instructions. Start of lesson Divide the classes into 6 groups: – Origin of replication – Repressor gene – Promoter – Multiple cloning.

Section H Cloning Vectors

Week 8. Outline Project Goal Extracting and biobricking KaiA and KaiB Synthesis update Western Blotting update Site-Specific Mutagenesis update Promoter.

Flashing Bacteria Morgan Haskell Coby Turner. What We Wanted To Do  University of California in San Diego  Biological synchronized clocks  Flash to.

CRISPR System Caroline Vrana Davidson College Synthetic Biology Summer 2012.

What are the Methods and Approaches Used to Identify and Study Arabidopsis Seed Knock- Out Mutations? Eric Newton Garen Polatoglu Rena Schweizer.

Recombinant DNA Technology

Chapter 20 DNA Transformation A. P. Biology Mr. Knowles Liberty Senior High School.

Pglo experiment What is ampicillin? A cell wall inhibiting antibiotic What happens to normal bacteria that are grown on agar plates with ampicillin in.

90- How can we make more insulin? V How can we make more insulin? By Transforming Bacteria V

BioSketchHarvard iGEM BioBrick: Aggregate Parts Quad Part Inverter (QPI) RBS CodingRegion Terminator Promoter RBSTerminator ReporterCodingRegion.

Fundamental Concepts in Genetic Logic Design - Principles of Control, Digital and Analog Devices Medicine 2B 徐廷儀 2009/03/11.

BioSketch The bacterial sketch pad. Hing Eng, Yves Wang, Jennifer Gao, Yin Li, Chris Doucette, Thomas Noriega Group Meeting

DNA Science Day 2 Extracting, Ligating and Transforming Physical Biology Bootcamp October 2006 Caltech.

DNA Science Day 2 Extracting, Ligating and Transforming APh162 Winter 2005 Caltech.

Genetic Jigsaw Class instructions. Start of lesson Divide the classes into 6 groups: – Origin of replication – Repressor gene – Promoter – Multiple cloning.

Tina Khoury Jeremy Gerbig Derek Blanchard Kerwin Dunham.

IGEM Arsenic Bioremediation Possibly finished biobrick for ArsR by adding a RBS and terminator. Will send for sequencing today or Monday.

Pierce College, Woodland Hills

In vivo gene cloning.

Section H Cloning Vectors.

Big Idea 3 – Investigation (Lab) 8. Recall how a gene of interest is obtained (PCR), inserted into a plasmid using restriction enzymes / DNA ligase, and.

Austin Jones Jace Dolphin. Methylosinus trichosporium culture courtesy of Dr. Alan DiSpirito, ISU Phenol/Chloroform Genomic DNA Extraction from protocol.

Introduction to pGLO lab Bacteria Transformation Please take these notes carefully. You do not need to write anything in RED.

By Kyle Albert & Josh Mixdorf. Why  Uncontrolled levels of estrogen in waste water treatment plants is a growing problem.  The high levels of estrogen.

Week 7 Wednesday: –Screening of library transformants –Innoculation of colonies for plasmid preps –Practice PCR Turn in Lab #11 Thursday: –Plasmid minipreps.

IGEM Bioware Arsenic PCR - 62% Successful! We're still having trouble with LamB and ArsB. Ligations and Transformations have been unsuccessful.

Brett Fuller Chase Meusel Holly Tjaden. -A neurotoxin produced by many organisms in nature -Causes paralysis in the victim -100 times more poisonous than.

Construction of a genetic toggle switch in Escherichia coli Farah and Tom.

Lab 20 Goals and Objectives: Exercise 58: Bacterial Counts on Foods

Restriction Analysis of pDRK and pGRN Original Plasmids

Restriction Enzyme Vector Ligase Enzyme Recombinant DNA DNA Construct Digestion ligation.

E. Coli Fluorescing Red Using Cold Temperature Sensor Prm + I B0032 Team: E Cool I Tina Khoury Jeremy Gerbig Kerwin Dunham Derek Blanchard.

Page 368 – 374. Gene Cloning gene cloning – Clones are used to examine the structure and function of the gene as well as the products of specific genes.

Treatments that stimulate the E.coli. to take up foreign plasmids include: 1.CaCL2 treatment 2.Heat shock 3.Incubation with ECORI 4.1 and and 3 6.All.

Genetic Engineering and Gene Regulation. What is genetic engineering? Manually adding new DNA to an organism, giving an organism traits it wouldn’t naturally.

Genetic Transformation of Bacteria and Gene Regulation.

Quorum Sensing iGEM2007. Target (eg Nickel) luxI OHHL luxR.

Cloning Vectors Enable DNA molecules to be replicated inside host (e.g., bacteria) cells. Features: 1. Origin of replication (ORI) 2. Cloning sites =

GENETIC TOGGLE SWITCH IN E. COLI COLLINS LAB (2000) Allen Lin 1.

Brown iGEM international genetically engineered machines competition August Update 1/55.

BioSketch The bacterial sketch pad. Yves Wang, Jennifer Gao, Yin Li Chris Doucette, Thomas Noriega, Hing Eng Group Meeting

CLONING VECTORS Shumaila Azam. IMPORTANT CLONING VECTORS.

From Genes to Genomes: Concepts and Applications of DNA Technology, Jeremy W. Dale, Malcolm von Schantz and Nick Plant. © 2012 John Wiley & Sons, Ltd.

BioWire Progress Report Week Six “Halfway There” Orr Ashenberg, Patrick Bradley, Connie Cheng, Kang-Xing Jin, Danny Popper, Sasha Rush.

Oxidative Burst Doug Tischer 1. General Idea Signal Galactose oxidase Catalase Buffers the cell from any leaky GOase expression = H 2 O 2 = H 2 O = O.

BioWire Progress Report Week Six “Halfway There” Orr Ashenberg, Patrick Bradley, Connie Cheng, Kang-Xing Jin, Danny Popper, Sasha Rush.

BioWire Progress Report Week Five Orr Ashenberg, Patrick Bradley, Connie Cheng, Kang-Xing Jin, Danny Popper, Sasha Rush.

Gene cloning  Steps  Preparation of insert  Modification  Ligation  Transformation.

Genetics of Bacteria Bacterial genome =.

Fac. of Agriculture, Assiut Univ.

Chapter 4 Recombinant DNA Technology

Exam 2 M 10/29 at 7-8:30pm in UTC 2.102A Review Th 10/25 at 5-7pm in WRW 102 Bonus #1 due now Last day to drop with a ‘Q’…10/24.

Methods of transformation

Genetic Transformation of Bacteria and Gene Regulation

Week 6: Electro-transformation and screening of the genomic library

iGEM Meeting #4 07/16/08 Presentation by Robert Ovadia

Example construct we would want to construct in B.subtilis

BioSketch The bacterial sketch pad.

Standardized and Scalable DNA Assembly

Week 1: Tutorial Outline

The final constructs, in theory

Virginia iGEM Workshop #2 High School Education Series.

Quorum Sensing Week 5 Plans.

OmpA C-Terminus Library

iGEM Meeting #3 07/09/08 Presentation by Robert Ovadia

1st Individually brainstorm and write down everything you remember from the Constitution! People Vocabulary Places Documents Constitution.

Cyanobacteria Oscillator

Presentation transcript:

The Goal Put together the parts we have now (and some parts we’re still missing) to make: LacI, TetR, and cI Lambda coding regions w/ hi/low constitutive promoters. GFP reporters with pLac, pTet, and pLambda. Put those two together on a p15a vector. Prepare new vectors with Biobrick sites and Duet vector origin(s), like CloDF13 (P13).

How do we get there? Hi/lo constitutive promoter + RBS + Repressor + Terminator Lac/ Tet/ Lambda Promoter + RBS + GFP + Terminator

Hi/lo constitutive repressor: the plan…

…and what we have so far

More on Last Week’s Progress We ligated RBS (P40) with TetR (P43) and cI Lambda (P44) coding regions to produce P56-7. Transformations were successful. A Biobrick part with RBS + lacI + terminators was transformed and miniprepped last week (P54), so that is now also available. P56-7 were digested, along with P38-9 (High/low constitutive promoters), and will be ligated and transformed today. NEXT: Add terminator(s) (P41, 61-4). Then insert into p15a vector w/ GFP reporter.

Plan for p15A Vector w/ GFP Reporter

What do we have so far? Everything but… Ligation of P45 + P18 happens today.

More on Last Week’s Progress We electroporated and transformed Shewie MR-1 with P58 and P59, as well as P12 (pET-Duet) and some of Remy’s plasmids to see if the pSC101 origin works in S1. P58 (Tet + GFP), as of Saturday, did not successfully transform. P59 plate grew, and colonies were GREEN under the microscope. =) Check those other plates! NEXT: Grow liquid cultures and miniprep/ make glycerol stocks. Troubleshoot as to why P58 didn’t grow. Work on P74 and P75.

Making a New Vector w/ CloDF13 ori

Progress on CloDF13 vector? We will ligate and transform modified P13 (CloDF13 ori) + P1 today.