Biotechnology (Chapter 20) Objectives Understand the background science behind the technology applications Understand the tools and details of the technology Develop familiarity with performing the select techniques
Plan Explaining the universality of the genetic material that makes DNA technology possible Brief explanation of use of bacteria in biotechnology applications Technical o Genetic variability in bacteria o Plasmids and plasmid vectors o Restriction Enzymes o Small volumes and proper use of micropipettes o Handling small Eppendorf tubes o Microfuges Overview of Biotechnology
DNA Technology & Biotechnology Background Information Definitions of biotechnology, DNA technology, genetic engineering Basic tools of biotechnology: genomic DNA, plasmid vectors, DNA modification enzymes (restriction enzymes) Basic Techniques: DNA gel electrophoresis, transformation, Polymerase Chain Reaction (PCR) Basic Lab Skills (Restriction Enzymes) Plasmid vector components & map Use of micropipette Setting-up restriction enzyme digests of a plasmid
Background Information Plasmids Source, composition & architecture Genes found on plasmids Functional DNA sequences of plasmids: o origin of replication o selectable marker- antibiotic resistance gene of interest o Gene(s) to be cloned o Control elements for gene expression Plasmid map Restriction Enzymes Where are they naturally found? Biological function Enzymatic activity- catalysis of sequence-specific cutting of DNA Staggered cutting-sticky ends Blunt cutting-no sticky ends Plasmid Maps with Restriction Enzyme sites Size of plasmid Position of restriction enzymes sites Size of fragments when cut and genes they carry
Genetic Material in Bacteria Chromosomal DNA circular Essential genes Extra-chromosomal small circular pieces of DNA- plasmids Non-essential useful genes
Plasmids Found in most bacteria, fungi, and plants but not in animal cells Multiply independently of bacterial chromosome Carry useful but non-essential genes, extend the biochemical capabilities of bacteria (antibiotic resistance genes) http://www.dnalc.org/resources/3d/20-mechanism-ofrecombination.html
DNA Modification enzymes in Restriction enzymes Endonucleases Bacteria Cleave DNA in a sequence-specific manner Different restriction enzymes cut different sequences Tools for cutting DNA during genetic engineering
Definitions Technology: the practical application of science to commerce or industry Biotechnology: the science of using living cells, and components of living cells, to produce goods and services.
Name of Bacteria Name of restriction Enzyme Restriction site Escherichia coli EcoR I 5' GAATTC 3' 3' CTTAAG 5' Haemophilus influenzae Serratia marscens Hind III Sma I 5' AAGCTT 3' 3' TTCGAA 5' 5' CCCGGG 3' 3' GGGCCC 5' Restriction enzymes http://highered.mcgrawhill.com/sites/0072437316/student_view0/chapter16/animations.html#
Definitions Engineering: exploiting basic mechanical principles to develop useful tools and objects Genetic engineering: Developing useful tools using genetic material (DNA)
Science behind DNA Technology Applications (Lecture topics) Basic understanding of the components of molecular genetics - DNA replication - Transcription - Translation - Control systems regulating all these processes
Basic tools of DNA technology - Genomic DNA - Vectors as shuttles for genes between organisms: Plasmids or viruses - Enzymes to manipulate DNA: ligase, DNA polymerase, & Restriction enzymes - Gel electrophoresis
Use of plasmids and restriction enzymes in cloning of genes Useful links http://www.sumanasinc.com/webcontent/animations/content/plasmid cloning.html http://www.wiley.com/legacy/college/boyer/0470003790/animations/ cloning/cloning.htm
Biotechnology uses DNA technology to produce useful products