Plasmid showing the operon for ampicilin resistance (ori) and the gene for ampicillin resistance (amp R )

Save this PDF as:
 WORD  PNG  TXT  JPG

Size: px
Start display at page:

Download "Plasmid showing the operon for ampicilin resistance (ori) and the gene for ampicillin resistance (amp R )"

Transcription

1 AP Biology Name AP Lab 8: Biotechnology (Bacterial Transformation) The bacterium Escherichia coli (E. coli) is a common inhabitant of the human colon and can be easily grown in inexpensive suspension culture. E. coli contains only around 5 million DNA base pairs (1/600 th the size of the human genome) located within a plasmid and can be easily manipulated in a lab. For these reasons, and others, E. coli is often used for molecular biology labs and labs testing the translation of enzymes from sequences of DNA. Bacteria transfer genes amongst themselves in three different ways. Conjugation, which is when genetic material is transferred from one bacterium to another of a different mating type; Transduction, which is when a virus acts as a vector (carrier) to transfer small pieces of DNA from one bacterium to another; and transformation, which involves a cell directly uptaking DNA from the environment into its own genome. Although effective, transformation only occurs at the end of logarithmic growth of a single parental strain. In a lab setting, restriction enzymes are used to insert and remove specific sections of DNA from plasmids. In this lab, you will use the concept of transformation to insert genes for antibiotic resistance and UV glowing into E. coli cells and observe whether translation occurs for these genes. Plasmid showing the operon for ampicilin resistance (ori) and the gene for ampicillin resistance (amp R )

2

3 Procedure In this lab, you will transform a gene for resistance to an antibiotic called ampicillin and another gene called pglo, which creates a protein that glows under UV light. Ampicillin is lethal to most strains of E. coli, so the gene should allow E. Coli to grow even in the presence of ampicillin. You will then observe the growth of E. coli in the presence of ampicillin and UV lights to see if the transformation was successful. DAY 1 1. Obtain one micro test tube labeled + for pglo and another - for no pglo. Place them in the foam tube rack. 2. Open the tubes and using a sterile transfer pipet, transfer 250 µl of ice cold 0.05 M transformation solution (CaCl 2 ). The CaCl 2 opens the plasma membrane of the E. coli. 3. Place the tubes on ice. 4. Use a sterile loop to pick up a single colony of bacteria from your starter plate. Pick up the +pglo tube and immerse the loop into the transformation solution at the bottom of the tube. Spin the loop between your index finger and thumb until the entire colony is dispersed in the transformation solution (with no floating chunks). Place the tube back in the tube rack in the ice. 5. Using a new sterile loop, repeat step 4 for the pglo tube. Try to get a colony of approximate similar size to each tube. 6. Examine the pglo plasmid DNA solution with the UV lamp. Note your observations. Immerse a new sterile loop into the plasmid DNA stock tube. Withdraw a loopful. There should be a film of plasmid solution across the ring. This is similar to seeing a soapy film across a ring for blowing soap bubbles. Mix the loopful into the cell suspension of the +pglo tube. 7. Close the tube and return it to the rack on ice. Also close the pglo tube. Do not add plasmid DNA to the pglo tube. 8. Incubate the tubes on ice for 10 minutes. Make sure to push the tubes all the way down in the rack so the bottom of the tubes stick out and make contact with the ice. 9. While the tubes are on ice, label your four agar plates on the bottom (not the lid) as follows a. LB/amp + pglo b. LB/amp/ara + pglo c. LB/amp pglo d. LB pglo i. *amp stands for Ampicillin. These plates have agar that has been laced with ampicillin. ii. **ara stands for Arabinose. Arabinose is a sugar that serves as a promoter for the pglo operon. 10. The heat shock step. Using the foam rack as a holder, transfer both the +pglo and pglo tubes into the water bath, set at 42 C, for exactly 50 seconds. Make sure to push the tubes all the way down in the rack so the bottom of the tubes stick out and make contact with the warm water. When the 50 seconds are done, place both tubes back on ice. For the best transformation results the change form the ice to 42 C and then back to the ice must be rapid.

4 a. Heat-shocking is essential because it will increase the permeability of the cell membrane, allowing the plasmid to enter the E. coli cell by creating a vacuum. 11. Incubate tubes on ice for 2 minutes. This will give the cells enough time to read the new ampicillin-resistance and pglo genes (if they have it) and translate it into a protein to hydrolyze ampicillin and build GFP, the gene from the pglo. 12. Remove the rack from the ice and place on the table. Open a tube and, using a new sterile pipet, add 250 µl of LB nutrient broth to the +pglo tube and close it. Mix by tapping with your finger. Repeat with a new sterile pipet for the pglo tube. Incubate at room temperature for 10 minutes. a. These 10 minutes are essential because it gives the E. coli cells time to transcribe and translate the ampicillin-hydrolyzing proteins and the pglo operon proteins. 13. Using a new sterile pipet for each tube, pipet 100 µl of the suspensions into their appropriate plates according to your labels. 14. Immediately spread the cells evenly by using a new sterile spreading rod for each plate. 15. Allow plates to set for several minutes. Then, stack your plates bottom-side up and tape them together. Put your group name and class period on the side facing up and place the stack in the 37 incubator. 16. Fill in the first column in Table 8.1. DAY When time to observe the cells, pull out of the incubator but DO NOT OPEN THE PLATES. Count the number of individual colonies and record in Table 8.1. Use a marker to mark each colony as it is counted. If the cell growth is too dense to count individual colonies, record lawn. 18. Complete the transformation efficiency analysis in Table 8.2. Post-Lab Questions 1. Indicate how each plate actually appeared compared to your prediction. 2. Which combination of plates best shows the transformation efficiency? Explain your reasoning. 3. Name two factors that influenced transformation efficiency? Explain the effect of each. 4. What are restriction enzymes, and how do they work? 5. What is the function of restriction enzymes in nature? Lab Book Requirements 1. Date of Lab 2. Title of Lab 3. Objective of Lab 4. Manipulated and Responding variables 5. Three Control Variables 6. Hypothesis 7. Table Table Conclusion 10. What is the next experiment you could perform? 11. Answers to Post-Lab Questions

5 Table 8.1: Colony Count. Plate Prediction: # of Colonies and UV Glowing? Number of Colonies Glowing under UV Light? LB/amp +pglo LB/amp/ara +pglo LB/amp pglo LB pglo Table 8.2: Transformation Efficiency. Transformation efficiency is expressed as the number of antibiotic-resistant colonies per microgram of pamp. Because transformation is limited to only those cells that are competent, increasing the amount of plasmid used does not necessarily increase the probability that a cell will be transformed. A sample of competent cells is usually saturated with small amounts of plasmid and excess DNA may actually interfere with the transformation process. a. Determine the total mass of pglo used by multiplying 10 µl of pglo at a concentration of µg/µl. Total mass = volume x concentration b. Review your procedure and calculate the total volume of cell suspension prepared c. Calculate the fraction of the total cell suspension that was spread on the plate by dividing the volume pipetted onto the plate by total volume (B). d. Determine the mass of pglo in cell suspension by multiplying the total mass of pglo (A) and the fraction of suspension spread (C). e. Determine the number of colonies per mg of plasmid by dividing the number of colonies observed by the mass of the pglo spread (D). This is your transformation efficiency.

Student Manual. pglo Transformation

Student Manual. pglo Transformation Student Manual pglo Transformation Lesson 1 Introduction to Transformation In this lab you will perform a procedure known as genetic transformation. Remember that a gene is a piece of DNA which provides

More information

Genetic Transformation Part 1

Genetic Transformation Part 1 Genetic Transformation Part 1 The beginning of an exploration of genetic transformation and the influence of environment on gene expression. * CONTENTS 1 Objectives... 1 1.1 Experimental Goal... 1 1.2

More information

Rapid Colony Transformation of E. coli with Plasmid DNA

Rapid Colony Transformation of E. coli with Plasmid DNA Rapid Colony Transformation of E. coli with Plasmid DNA Introduction: The bacterium Escherichia coli (E. coli) is an ideal organism for the molecular geneticist to manipulate and has been used extensively

More information

pglo Bacterial Transformation Evaluation copy

pglo Bacterial Transformation Evaluation copy pg Bacterial Transformation Computer 6A Introduction to Transformation In this lab, you will perform a procedure known as genetic transformation. Genetic transformation literally means change caused by

More information

Green Fluorescent Protein (GFP): Genetic Transformation, Synthesis and Purification of the Recombinant Protein

Green Fluorescent Protein (GFP): Genetic Transformation, Synthesis and Purification of the Recombinant Protein Green Fluorescent Protein (GFP): Genetic Transformation, Synthesis and Purification of the Recombinant Protein INTRODUCTION Green Fluorescent Protein (GFP) is a novel protein produced by the bioluminescent

More information

LAB 16 Rapid Colony Transformation of E. coli with Plasmid DNA

LAB 16 Rapid Colony Transformation of E. coli with Plasmid DNA LAB 16 Rapid Colony Transformation of E. coli with Plasmid DNA Objective: In this laboratory investigation, plasmids containing fragments of foreign DNA will be used to transform Escherichia coli cells,

More information

Bacterial Transformation with Green Fluorescent Protein. Table of Contents Fall 2012

Bacterial Transformation with Green Fluorescent Protein. Table of Contents Fall 2012 Bacterial Transformation with Green Fluorescent Protein pglo Version Table of Contents Bacterial Transformation Introduction..1 Laboratory Exercise...3 Important Laboratory Practices 3 Protocol...... 4

More information

GENETIC TRANSFORMATION OF BACTERIA WITH THE GENE FOR GREEN FLUORESCENT PROTEIN (GFP)

GENETIC TRANSFORMATION OF BACTERIA WITH THE GENE FOR GREEN FLUORESCENT PROTEIN (GFP) GENETIC TRANSFORMATION OF BACTERIA WITH THE GENE FOR GREEN FLUORESCENT PROTEIN (GFP) LAB BAC3 Adapted from "Biotechnology Explorer pglo Bacterial Transformation Kit Instruction Manual". (Catalog No. 166-0003-EDU)

More information

Bacterial Transformation with Green Fluorescent Protein. pgfp Version. Table of Contents Fall 2012

Bacterial Transformation with Green Fluorescent Protein. pgfp Version. Table of Contents Fall 2012 Bacterial Transformation with Green Fluorescent Protein pgfp Version Table of Contents Bacterial Transformation Introduction..1 Laboratory Exercise...3 Important Laboratory Practices 3 Protocol...... 4

More information

pglo Bacterial Transformation

pglo Bacterial Transformation Introduction pglo Bacterial Transformation Biotechnology refers to technology used to manipulate DNA. The procedures are often referred to as genetic engineering. DNA is the genetic material of all living

More information

Transformation 1. Introduction: 221 Lab Manual.KCBurke

Transformation 1. Introduction: 221 Lab Manual.KCBurke Transformation 1 http://www.cdc.gov/drugresistance/about.html Antibiotic resistance is a critical problem in healthcare today. How do bacteria become resistant to antibiotics? In this lab we will focus

More information

Transformation of the bacterium E. coli. using a gene for Green Fluorescent Protein

Transformation of the bacterium E. coli. using a gene for Green Fluorescent Protein Transformation of the bacterium E. coli using a gene for Green Fluorescent Protein Background In molecular biology, transformation refers to a form of genetic exchange in which the genetic material carried

More information

AP BIOLOGY. Investigation #8 Biotechnology: Bacterial Transformation. Slide 1 / 31. Slide 2 / 31. Slide 3 / 31. Investigation #8: Transformation

AP BIOLOGY. Investigation #8 Biotechnology: Bacterial Transformation. Slide 1 / 31. Slide 2 / 31. Slide 3 / 31. Investigation #8: Transformation New Jersey Center for Teaching and Learning Slide 1 / 31 Progressive Science Initiative This material is made freely available at www.njctl.org and is intended for the non-commercial use of students and

More information

Transformation Kit BACTERIAL TRANSFORMATION: GREEN FLUORESCENT PROTEIN. Partnership for Biotechnology and Genomics Education

Transformation Kit BACTERIAL TRANSFORMATION: GREEN FLUORESCENT PROTEIN. Partnership for Biotechnology and Genomics Education Transformation Kit BACTERIAL TRANSFORMATION: GREEN FLUORESCENT PROTEIN Partnership for Biotechnology and Genomics Education Barbara Soots Linda Curro Education Coordinator University of California Davis

More information

Student Manual. pglo Transformation

Student Manual. pglo Transformation Student Manual pglo Transformation Lesson 1 Introduction to Transformation In this lab you will perform a procedure known as a genetic transformation. Remember that a gene is a piece of DNA which provides

More information

Lab Exercise: Transformation

Lab Exercise: Transformation Lab Exercise: Transformation Background Genetic transformation is used in many areas of biotechnology, and, at its heart, requires two things: Donor DNA and recipient cells. Cells which receive the donor

More information

Student Manual. pglo Transformation

Student Manual. pglo Transformation Student Manual pglo Transformation Lesson 1 Introduction to Transformation In this lab you will perform a procedure known as genetic transformation. Remember that a gene is a piece of DNA which provides

More information

The Chemical Structure of Ampicillin

The Chemical Structure of Ampicillin Bio 210A Bacterial Transformation-Gene Cloning Purpose To understand the relevance of gene cloning to the biotechnology industry. To understand the definition of transformation, clone, and cloning vector.

More information

DNA For those that downloaded the notes a reward! Paste the flowchart in your notebook. However, please read, study and understand the procedure.

DNA For those that downloaded the notes a reward! Paste the flowchart in your notebook. However, please read, study and understand the procedure. DNA For those that downloaded the notes a reward! Paste the flowchart in your notebook. However, please read, study and understand the procedure. Genetic Engineering Lecture Notes Bacteria contain genes

More information

Genetics and Information Transfer

Genetics and Information Transfer Genetics and Information Transfer Big Idea 3 investigation 8 BIOTECHNOLOGY: BACTERIAL TRANSFORMATION* How can we use genetic engineering techniques to manipulate heritable information? BACKGROUND Are genetically

More information

Lab 9: Bacterial Transformation with pglo

Lab 9: Bacterial Transformation with pglo Name: Lab 9: Bacterial Transformation with pglo OBJECTIVES: ο Practice formulating hypotheses, predictions, and experimental design. ο Describe the principles of bacterial transformation. ο Explain the

More information

MOLECULAR GENETICS GENETIC ENGINEERING RECOMBINANT DNA. Molecular Genetics Activity #6 page 1

MOLECULAR GENETICS GENETIC ENGINEERING RECOMBINANT DNA. Molecular Genetics Activity #6 page 1 AP BIOLOGY MOLECULAR GENETICS ACTIVITY #6 NAME DATE HOUR RECOMBINANT DNA GENETIC ENGINEERING Molecular Genetics Activity #6 page 1 GENETIC ENGINEERING Molecular Genetics Activity #6 page 2 PART I: PRODUCING

More information

Biology Lab Activity 4-5 DNA Transformation

Biology Lab Activity 4-5 DNA Transformation Biology Lab Activity 4-5 DNA Transformation Scientists can insert genes into bacteria. The genes inserted in the Indo-Blu process (this lab) are on a circular piece of DNA called a plasmid. (The plasmid

More information

Genetic transformation literally means change caused by genes.

Genetic transformation literally means change caused by genes. pglo Bacterial Transformation Practical What is transformation? Genetic transformation literally means change caused by genes. It occurs when a cell takes up (takes inside) and expresses a new piece of

More information

Lab 10: Bacterial Transformation, part 2, DNA plasmid preps, Determining DNA Concentration and Purity

Lab 10: Bacterial Transformation, part 2, DNA plasmid preps, Determining DNA Concentration and Purity Lab 10: Bacterial Transformation, part 2, DNA plasmid preps, Determining DNA Concentration and Purity Today you analyze the results of your bacterial transformation from last week and determine the efficiency

More information

Biotechnology Explorer

Biotechnology Explorer arac ori pglo bla GFP Biotechnology Explorer pglo Bacterial Transformation Kit Catalog Number 166-0003EDU explorer.bio-rad.com Store components of this kit at room temperature. Duplication of any part

More information

GFP Transformation Genetic Manipulations

GFP Transformation Genetic Manipulations MODULE 2 Objective 2.1 Lesson E GFP Transformation Genetic Manipulations Course Advanced Biotechnology Unit DNA Technology Essential Question How is foreign DNA genes taken up by organisms and expressed?

More information

pglo Transformation and Inquiry Kit A ThINQ! Investigation Student Manual

pglo Transformation and Inquiry Kit A ThINQ! Investigation Student Manual pglo Transformation and Inquiry Kit A ThINQ! Investigation Student Manual Dear Students Mice that glow fl uorescent green. Plants that turn red when grown near a land mine. Goats that make milk that can

More information

Evaluation copy. Easy Transformation of E.coli Using GPF. Computer 6A

Evaluation copy. Easy Transformation of E.coli Using GPF. Computer 6A Easy Transformation of E.coli Using GPF Computer 6A Introduction Transformation can be defined as the uptake and expression of foreign DNA by a living cell. In this laboratory, you will transform Escherichia

More information

Biotechnology Explorer

Biotechnology Explorer arac ori pglo bla GFP Biotechnology Explorer Bacterial Transformation The pglo System Catalog Number 166-0003-EDU www.bio-rad.com For Technical Service Call Your Local Bio-Rad Office or in the U.S. Call

More information

DNA Isolation from Strawberries Developed by Diane Sweeney

DNA Isolation from Strawberries Developed by Diane Sweeney Eileen Roach Biology This year my students did various biotechnology labs that helped them understand the concepts of diffusion through a membrane, polarity of molecules, DNA structure, and bacterial transformation.

More information

The Effects of Plasmid on Genotype and Phenotype (Revised 1/31/96) Introduction

The Effects of Plasmid on Genotype and Phenotype (Revised 1/31/96) Introduction The Effects of Plasmid on Genotype and Phenotype (Revised 1/31/96) Introduction Plasmids are small circular DNA molecules that often found in bacteria in addition to the large circular DNA molecule of

More information

Transformation of Competent Cells with a Recombinant Plasmid Carl Estrella, Merced College, Merced, CA

Transformation of Competent Cells with a Recombinant Plasmid Carl Estrella, Merced College, Merced, CA INTRODUCTION To close the yellow note, click once to select it and then click the box in the upper left corner. To open the note, double click (Mac OS) or right click (Windows) on the note icon. Transformation

More information

Transformation. Making Change Happen

Transformation. Making Change Happen Transformation Making Change Happen Genetic Engineering Definition: The alteration of an organism s genetic, or hereditary, material to eliminate undesirable characteristics or to produce desirable new

More information

AP BIOLOGY 2009 SCORING GUIDELINES (Form B)

AP BIOLOGY 2009 SCORING GUIDELINES (Form B) AP BIOLOGY 2009 SCORING GUIDELINES (Form B) Question 1 Describe how a plasmid can be genetically modified to include a piece of foreign DNA that alters the phenotype of bacterial cells transformed with

More information

Lab 9: Bacterial Transformation & Spectrophotometry, Part 1

Lab 9: Bacterial Transformation & Spectrophotometry, Part 1 Lab 9: Bacterial Transformation & Spectrophotometry, Part 1 Activity 9a Bacterial Transformation, Part 1 Purpose and Background In this lab, you will perform a procedure known as genetic transformation.

More information

Bacterial Transformation and Plasmid Purification. Chapter 5: Background

Bacterial Transformation and Plasmid Purification. Chapter 5: Background Bacterial Transformation and Plasmid Purification Chapter 5: Background History of Transformation and Plasmids Bacterial methods of DNA transfer Transformation: when bacteria take up DNA from their environment

More information

GROWING BACTERIA INTRODUCTION

GROWING BACTERIA INTRODUCTION GROWING BACTERIA INTRODUCTION E. coli is a normal part of the bacterial flora of the human gut. It is not generally considered pathogenic, although some strains are highly toxic (recent food poisonings

More information

Biotechnology Explorer

Biotechnology Explorer arac ori pglo bla GFP Biotechnology Explorer pglo Bacterial Transformation Kit Catalog #166-0003EDU explorer.bio-rad.com See individual components for storage temperature. Duplication of any part of this

More information

MOLEBIO LAB #6: Bacterial Culture Techniques Part I

MOLEBIO LAB #6: Bacterial Culture Techniques Part I MOLEBIO LAB #6: Bacterial Culture Techniques Part I Introduction: This lab introduces an introduction to plating and culturing E. coli on LB agar plates such that single cells can be isolated from one

More information

SAMPLE. Bacterial Transformation. Lab 8 BACKGROUND INFORMATION. Neo/SCI Student s Guide Name... Teacher/Section...

SAMPLE. Bacterial Transformation. Lab 8 BACKGROUND INFORMATION. Neo/SCI Student s Guide Name... Teacher/Section... 1431489 REV 001 Neo/SCI Lab 8 Bacterial Transformation BACKGROUND INFORMATION What Is Biotechnology? Before you start doing biotechnology laboratory exercises, it is important to know exactly what biotechnology

More information

Genetic Engineering: Transforming Bacteria

Genetic Engineering: Transforming Bacteria Genetic Engineering: Transforming Bacteria Introduction Lesson Introduction In this laboratory experiment, students will transform the phenotype of bacteria through the use of genetic engineering. A non

More information

DNA CAN BE TRANSFERRED BETWEEN BACTERIA GENETIC ENGINEERING USING RECOMBINANT DNA TECHNOLOGY

DNA CAN BE TRANSFERRED BETWEEN BACTERIA GENETIC ENGINEERING USING RECOMBINANT DNA TECHNOLOGY Bacterial Transformation DNA CAN BE TRANSFERRED BETWEEN BACTERIA Background Information Plasmid Transformed Cell Figure 1: Bacterial Transformation Quick Reference Abbreviations GFP pgfp gfp Green fl uorescent

More information

AP BIOLOGY 2007 SCORING GUIDELINES

AP BIOLOGY 2007 SCORING GUIDELINES AP BIOLOGY 2007 SCORING GUIDELINES Question 4 A bacterial plasmid is 100 kb in length. The plasmid DNA was digested to completion with two restriction enzymes in three separate treatments: EcoRI, HaeIII,

More information

GLOW-IN-THE-DARK BACTERIA (Transformation Lab)

GLOW-IN-THE-DARK BACTERIA (Transformation Lab) GLOW-IN-THE-DARK BACTERIA (Transformation Lab) A. BACKGROUND INFORMATION The Problem of Infectious Disease: At the turn of this century, infectious diseases like pneumonia and influenza caused high numbers

More information

Genetics and Information Transfer

Genetics and Information Transfer Genetics and Information Transfer Big Idea 3 investigation 8 BIOTECHNOLOGY: BACTERIAL TRANSFORMATION* How can we use genetic engineering techniques to manipulate heritable information? BACKGROUND Are genetically

More information

Molecular Biology. Lab #2 Growth of E. coli Bacteria

Molecular Biology. Lab #2 Growth of E. coli Bacteria Molecular Biology Name Lab #2 Growth of E. coli Bacteria Escherichia coli (E. coli) is a rod-shaped (bacillus), enteric (gut) bacterium of the family Enterobacteriaceae. It is a normal resident of the

More information

Cloning GFP into Mammalian cells

Cloning GFP into Mammalian cells Protocol for Cloning GFP into Mammalian cells Studiepraktik 2013 Molecular Biology and Molecular Medicine Aarhus University Produced by the instructors: Tobias Holm Bønnelykke, Rikke Mouridsen, Steffan

More information

Transformation Protocol

Transformation Protocol To make Glycerol Stocks of Plasmids ** To be done in the hood and use RNase/DNase free tips** 1. In a 10 ml sterile tube add 3 ml autoclaved LB broth and 1.5 ul antibiotic (@ 100 ug/ul) or 3 ul antibiotic

More information

Producing a Strain of E. coli that Glows in the Dark Pamela C. Edgerton, Governor s School for Government and International Studies, Richmond, VA

Producing a Strain of E. coli that Glows in the Dark Pamela C. Edgerton, Governor s School for Government and International Studies, Richmond, VA Producing a Strain of E. coli that Glows in the Dark Pamela C. Edgerton, Governor s School for Government and International Studies, Richmond, VA INTRODUCTION To close the yellow note, click once to select

More information

Transformation Ameer Effat M. Elfarash

Transformation Ameer Effat M. Elfarash Transformation Ameer Effat M. Elfarash Dept. of Genetics Fac. of Agriculture, Assiut Univ. amir_effat@yahoo.com DNA Cloning Overview Introduction to cell A. Cell type: Bacteria, Mammalian cells, insect

More information

Transformation of of E. coli with a Green Fluorescent Protein Plasmid

Transformation of of E. coli with a Green Fluorescent Protein Plasmid The Biotechnology Education Company Revised and Updated EDVO-Kit # 223 Transformation of of E. coli with a Green Fluorescent Protein Plasmid Storage: See Page 3 for specific storage instructions EXPERIMENT

More information

Isolation and Electrophoresis of Plasmid DNA

Isolation and Electrophoresis of Plasmid DNA Name Date Isolation and Electrophoresis of Plasmid DNA Prior to lab you should be able to: o Explain what cloning a gene accomplishes for a geneticist. o Describe what a plasmid is. o Describe the function

More information

Agrobacterium tumefaciens-mediated transformation of Colletotrichum graminicola and Colletotrichum sublineolum

Agrobacterium tumefaciens-mediated transformation of Colletotrichum graminicola and Colletotrichum sublineolum Agrobacterium tumefaciens-mediated transformation of Colletotrichum graminicola and Colletotrichum sublineolum Flowers and Vaillancourt, 2005. Current Genetics 48: 380-388 NOTE added by L. Vaillancourt:

More information

AP Biology -- John Burroughs School -- M. Bahe

AP Biology -- John Burroughs School -- M. Bahe Objectives for Test Eight: Chapter 13 14, 15.2, 17.1 DNA, Protein Synthesis, Gene Regulation & Biotechnology You should be able to: 1. Identify the scientists who contributed pieces of the genetic puzzle,"

More information

Plasmid Isolation. Prepared by Latifa Aljebali Office: Building 5, 3 rd floor, 5T250

Plasmid Isolation. Prepared by Latifa Aljebali Office: Building 5, 3 rd floor, 5T250 Plasmid Isolation Prepared by Latifa Aljebali Office: Building 5, 3 rd floor, 5T250 Plasmid Plasmids are small, double strand, closed circular DNA molecules. Isolated from bacterial cells. Replicate independently

More information

Recombinant DNA and Biotechnology

Recombinant DNA and Biotechnology Recombinant DNA and Biotechnology Chapter 18 Lecture Objectives What Is Recombinant DNA? How Are New Genes Inserted into Cells? What Sources of DNA Are Used in Cloning? What Other Tools Are Used to Study

More information

Activity #5b. Plasmid DNA Isolation, Restriction Enzyme Digestion & Bacterial Transformation

Activity #5b. Plasmid DNA Isolation, Restriction Enzyme Digestion & Bacterial Transformation Activity #5b. Plasmid DNA Isolation, Restriction Enzyme Digestion & Bacterial Transformation Learning Goals: To learn about bacterial plasmids, one of the basic tools of genetic engineering To purify plasmid

More information

The ramylase Project by Ellyn Daugherty

The ramylase Project by Ellyn Daugherty PR078 G-Biosciences 1-800-628-7730 1-314-991-6034 technical@gbiosciences.com A Geno Technology, Inc. (USA) brand name The ramylase Project by Ellyn Daugherty Transformation of E. coli with pamylase (Lab

More information

Welcome to Implementing Inquirybased Microbial Project. Veronica Ardi, PhD

Welcome to Implementing Inquirybased Microbial Project. Veronica Ardi, PhD Welcome to Implementing Inquirybased Microbial Project Veronica Ardi, PhD Microbiology Laboratory Courses CourseSmart: ebook resources http://instructors.coursesmart.com/ Microbiology Laboratory Courses

More information

Gene Cloning Technology

Gene Cloning Technology Gene Cloning Technology Also known as: Genetic engineering or Genetic manipulation (GM) technology implies precision engineering being applied to DNA molecules Recombinant DNA technology - implies that

More information

Zback Faster Ligation Kit

Zback Faster Ligation Kit Zback Faster Ligation Kit For the highest efficiency cloning of PCR products either blunt or sticky-end Kit Contents Contents TGVTB04 TGVTB04-2 pzback/blunt vector 10 µl 20 µl T4 DNA Ligase 5 µl 10 µl

More information

Recombinant Paper Plasmids Cut-and-Paste Biotechnology

Recombinant Paper Plasmids Cut-and-Paste Biotechnology Recombinant Paper Plasmids Cut-and-Paste Biotechnology OBJECTIVE / RIONALE Bioengineers make news using recombinant DNA techniques in hopes of curing genetic diseases, better understanding cancer, and

More information

TRANSFORMATION OF BACTERIA WITH DIFFERENT PLASMIDS. Objectives

TRANSFORMATION OF BACTERIA WITH DIFFERENT PLASMIDS. Objectives TRANSFORMATION OF BACTERIA WITH DIFFERENT PLASMIDS Objectives To understand the concept of DNA as genetic material through the process of transformation. To test the conditions that make cells competent

More information

Genetic Engineering. Take part in research to combat atherosclerosis. Experiment Workshop PROTOCOL

Genetic Engineering. Take part in research to combat atherosclerosis. Experiment Workshop PROTOCOL Genetic Engineering Take part in research to combat atherosclerosis Experiment Workshop PROTOCOL Genetic Engineering Searching for a target for the treatment of atherosclerosis Introduction Biomedical

More information

Recombinant DNA technology (genetic engineering) involves combining genes from different sources into new cells that can express the genes.

Recombinant DNA technology (genetic engineering) involves combining genes from different sources into new cells that can express the genes. Recombinant DNA technology (genetic engineering) involves combining genes from different sources into new cells that can express the genes. Recombinant DNA technology has had-and will havemany important

More information

LAB 7 DNA RESTRICTION for CLONING

LAB 7 DNA RESTRICTION for CLONING BIOTECHNOLOGY I DNA RESTRICTION FOR CLONING LAB 7 DNA RESTRICTION for CLONING STUDENT GUIDE GOALS The goals of this lab are to provide the biotech student with experience in DNA digestion with restriction

More information

Transferring a Broth Culture to Fresh Broth

Transferring a Broth Culture to Fresh Broth Sterile Technique It is very important in microbiology to work with pure cultures. Unfortunately this is difficult. The world around us is covered with microorganisms. Microorganisms are even carried on

More information

Effects of Antibiotics on Bacterial Growth and Protein Synthesis: Student Laboratory Manual

Effects of Antibiotics on Bacterial Growth and Protein Synthesis: Student Laboratory Manual Effects of Antibiotics on Bacterial Growth and Protein Synthesis: Student Laboratory Manual I. Purpose...1 II. Introduction...1 III. Inhibition of Bacterial Growth Protocol...2 IV. Inhibition of in vitro

More information

30. Genetics and recombination in bacteria Lecture Outline 11/16/05. The Bacterial Genome and Its Replication The bacterial chromosome

30. Genetics and recombination in bacteria Lecture Outline 11/16/05. The Bacterial Genome and Its Replication The bacterial chromosome 30. Genetics and recombination in bacteria Lecture Outline 11/16/05 Replication in bacteria Types of recombination in bacteria Transduction by phage Conjugation ( mating ) F+ plasmids Hfr s Transformation

More information

A and B are not absolutely linked. They could be far enough apart on the chromosome that they assort independently.

A and B are not absolutely linked. They could be far enough apart on the chromosome that they assort independently. Name Section 7.014 Problem Set 5 Please print out this problem set and record your answers on the printed copy. Answers to this problem set are to be turned in to the box outside 68-120 by 5:00pm on Friday

More information

Transformation of E.coli with pgal

Transformation of E.coli with pgal The Biotechnology Education Company ATTENTION! This experiment includes either BactoBeads or LyphoCells. If you have received LyphoCells, please refer to the addendum posted on the last page of this literature.

More information

Biotechnology and Recombinant DNA (Chapter 9) Lecture Materials for Amy Warenda Czura, Ph.D. Suffolk County Community College

Biotechnology and Recombinant DNA (Chapter 9) Lecture Materials for Amy Warenda Czura, Ph.D. Suffolk County Community College Biotechnology and Recombinant DNA (Chapter 9) Lecture Materials for Amy Warenda Czura, Ph.D. Suffolk County Community College Primary Source for figures and content: Eastern Campus Tortora, G.J. Microbiology

More information

Chapter 20: Biotechnology: DNA Technology & Genomics

Chapter 20: Biotechnology: DNA Technology & Genomics Biotechnology Chapter 20: Biotechnology: DNA Technology & Genomics The BIG Questions How can we use our knowledge of DNA to: o Diagnose disease or defect? o Cure disease or defect? o Change/improve organisms?

More information

Lecture 36: Basics of DNA Cloning-II

Lecture 36: Basics of DNA Cloning-II Lecture 36: Basics of DNA Cloning-II Note: Before starting this lecture students should have completed Lecture 35 Sequential steps involved in DNA cloning using plasmid DNA as vector: Molecular cloning

More information

The Biotechnology Education Company. Transformation with Green Fluorescent Protein (GFP) Storage: See Page 3 for specific storage instructions

The Biotechnology Education Company. Transformation with Green Fluorescent Protein (GFP) Storage: See Page 3 for specific storage instructions The Biotechnology Education Company Revised and Updated EDVO-Kit # 223/AP08 Transformation with Green Fluorescent Protein (GFP) Storage: See Page 3 for specific storage instructions EXPERIMENT OBJECTIVE:

More information

Biotechnology Explorer

Biotechnology Explorer Biotechnology Explorer Green Fluorescent Protein (GFP) Purification Kit Catalog Number 166-0005-EDU www.bio-rad.com For Technical Service Call Your Local Bio-Rad Office or in the U.S. Call 1-800-4BIORAD

More information

In order to be useful, a smear must have the following qualities:

In order to be useful, a smear must have the following qualities: Smear Preparation and Simple Stain Objectives: Make bacterial smear slides (usually called smears) Distinguish cells on these slides using a simple stain procedure Unstained microbial cells are nearly

More information

TransformAid Bacterial Transformation Kit

TransformAid Bacterial Transformation Kit Home Contacts Order Catalog Support Search Alphabetical Index Numerical Index Restriction Endonucleases Modifying Enzymes PCR Kits Markers Nucleic Acids Nucleotides & Oligonucleotides Media Transfection

More information

AP Biology Review Packet 4: Viruses, Bacteria and Expression & DNA Technology

AP Biology Review Packet 4: Viruses, Bacteria and Expression & DNA Technology AP Biology Review Packet 4: Viruses, Bacteria and Expression & DNA Technology 3A1- DNA, and in some cases RNA, is the primary source of heritable information. 3B1- Gene Regulation results in differential

More information

Biological Sciences Initiative

Biological Sciences Initiative Biological Sciences Initiative HHMI Student Activities Measuring Antibiotic Resistance Introduction: You might be aware that antibiotics were once thought of as a magic bullet; a nearly perfect drug for

More information

BACTERIAL ENUMERATION

BACTERIAL ENUMERATION BACTERIAL ENUMERATION In the study of microbiology, there are numerous occasions when it is necessary to either estimate or determine the number of bacterial cells in a broth culture or liquid medium.

More information

MGC premier full length cdna and ORF clones

MGC premier full length cdna and ORF clones MGC premier full length cdna and ORF clones TCH1003, TCM1004, TCR1005, TCB1006, TCL1007, TCT1008, TCZ1009, TOH6003, TOM6004, TOZ6009, TCHS1003, TCMS1004, TCRS1005, TCBS1006, TCLS1007, TCTS1008 MGC premier

More information

Biotechnology Explorer TM. Green Fluorescent Protein (GFP) Purification Kit. Instruction Manual

Biotechnology Explorer TM. Green Fluorescent Protein (GFP) Purification Kit. Instruction Manual Biotechnology Explorer TM Green Fluorescent Protein (GFP) Purification Kit Instruction Manual Catalog #166-0005EDU explorer.bio-rad.com Duplication of any part of this document is permitted for classroom

More information

Transformation of E.coli with pgal

Transformation of E.coli with pgal The Biotechnology Education Company Revised and Updated EDVO-Kit # 221 Transformation of E.coli with pgal Storage: See Page 3 for specific storage instructions EXPERIMENT OBJECTIVE: The objective of this

More information

Chapter 12 - DNA Technology

Chapter 12 - DNA Technology Bio 100 DNA Technology 1 Chapter 12 - DNA Technology Among bacteria, there are 3 mechanisms for transferring genes from one cell to another cell: transformation, transduction, and conjugation 1. Transformation

More information

Dilution Theory and Problems

Dilution Theory and Problems Dilution Theory and Problems Microorganisms are often counted in the laboratory using such methods as the viable plate count where a dilution of a sample is plated onto (or into) an agar medium. After

More information

Development of M13 as a Single-stranded Cloning Vector: Insertion of the Tn3 Transposon into the Genome of M13

Development of M13 as a Single-stranded Cloning Vector: Insertion of the Tn3 Transposon into the Genome of M13 Development of M13 as a Single-stranded Cloning Vector: Insertion of the Tn3 Transposon into the Genome of M13 Dan S. Ray and Karin Kook Molecular Biology Institute and Department of Biology University

More information

Chapter 9. Biotechnology and Recombinant DNA Biotechnology and Recombinant DNA

Chapter 9. Biotechnology and Recombinant DNA Biotechnology and Recombinant DNA Chapter 9 Biotechnology and Recombinant DNA Biotechnology and Recombinant DNA Q&A Interferons are species specific, so that interferons to be used in humans must be produced in human cells. Can you think

More information

Microbiology Laboratory: MOLECULAR IDENTIFICATION OF UNKNOWN BACTERIA

Microbiology Laboratory: MOLECULAR IDENTIFICATION OF UNKNOWN BACTERIA Microbiology Laboratory: MOLECULAR IDENTIFICATION OF UNKNOWN BACTERIA Classical Microbiology courses are typically structured to introduce the identification of bacterial species using a series of biochemical

More information

Many cells will not take up plasmid during transformation Cells with plasmid can be identified because original plasmid contained gene for antibiotic

Many cells will not take up plasmid during transformation Cells with plasmid can be identified because original plasmid contained gene for antibiotic Many cells will not take up plasmid during transformation Cells with plasmid can be identified because original plasmid contained gene for antibiotic resistance (ampicillin) Use medium with ampicillin

More information

Biotechnology: Bacterial Transformation with Green Fluorescent Protein

Biotechnology: Bacterial Transformation with Green Fluorescent Protein BIG IDEA 3 8 EDVO-Kit: Biotechnology: Bacterial Transformation with Green Fluorescent Protein See Page 3 for storage instructions. EXPERIMENT OBJECTIVE: The objective of this experiment is to develop an

More information

ANTIBIOTIC RESISTANCE & BACTERIAL TRANSFORMATION

ANTIBIOTIC RESISTANCE & BACTERIAL TRANSFORMATION ANTIBIOTIC RESISTANCE & BACTERIAL TRANSFORMATION STUDENT MANUAL Robin Ulep, Rebecca Sanchez Pitre, Stephanie Messina & Jawed Alam ABOUT BEST SCIENCE! Acknowledgements: The Antibiotic Resistance & Bacterial

More information

The E. coli Insulin Factory

The E. coli Insulin Factory The E. coli Insulin Factory BACKGROUND Bacteria have not only their normal DNA, they also have pieces of circular DNA called plasmids. Plasmids are a wonderfully ally for biologists who desire to get bacteria

More information

Growth of E. coli and determination of its doubling times on enriched and minimal media.

Growth of E. coli and determination of its doubling times on enriched and minimal media. Period 4 (Feb. 12) Growth of E. coli and determination of its doubling times on enriched and minimal media. The study of the growth of bacterial cultures does not constitute a specialized subject or branch

More information

Chapter 10 Manipulating Genes

Chapter 10 Manipulating Genes How DNA Molecules Are Analyzed Chapter 10 Manipulating Genes Until the development of recombinant DNA techniques, crucial clues for understanding how cell works remained lock in the genome. Important advances

More information

LAB 9 RECOMBINANT DNA LIGATION

LAB 9 RECOMBINANT DNA LIGATION BIOECHNOLOGY I RECOMBINN DN LIGION LB 9 RECOMBINN DN LIGION SUDEN GUIDE GOL he objective of this lab is to perform DN ligation to construct a recombinant plasmid. OBJECIVES fter completion, the student

More information

Today-applications: Medicine-better health Pharmaceutical-production of antibiotics Foods-wine, cheese, beer Agriculture-selective breeding

Today-applications: Medicine-better health Pharmaceutical-production of antibiotics Foods-wine, cheese, beer Agriculture-selective breeding I. Genetic Engineering modification of DNA of organisms to produce new genes with new characteristics -genes are small compared to chromosomes -need methods to get gene-sized pieces of DNA -direct manipulation

More information

UTILIZATION of PLASMA ACTIVATED WATER in Biotechnology, Pharmacology and Medicine. JSC TECHNOSYSTEM-ECO Moscow, Russia April, 2009

UTILIZATION of PLASMA ACTIVATED WATER in Biotechnology, Pharmacology and Medicine. JSC TECHNOSYSTEM-ECO Moscow, Russia April, 2009 UTILIZATION of PLASMA ACTIVATED WATER in Biotechnology, Pharmacology and Medicine JSC TECHNOSYSTEM-ECO Moscow, Russia April, 2009 METHOD of WATER ACTIVATION with PLASMA of GAS DISCHARGE ANODE VACUUM WATER

More information