1. C HAPTER 4: I NTRODUCTION TO S TUDYING DNA Introduction to Biotechnology, BIOL1414 Austin Community College, Biotechnology Dept

2. L EARNING O UTCOMES Describe the structure and function of DNA and explain the process by which it encodes for proteins Differentiate between eukaryotic and prokaryotic chromosomal structure and explain how this difference impacts gene regulation in the two cell types Differentiate between bacterial cultures grown in liquid and solid media and explain how to prepare each media type using sterile technique Discuss the characteristics of viruses and their importance in genetic engineering Explain the fundamental process of genetic engineering and give examples of the following applications: recombinant DNA technology, site-specific mutagenesis, and gene therapy Describe the process of gel electrophoresis and explain how the characteristics of molecules affect their migration through a gel Note about this PowerPoint – There are several links in this PPT that allow you to explore more into different topics. Some of these links are animations, movies, or exercises. Please note, you must be in the slide show to activate the links. You can press F5 any time to active the slide show and “Esc” to exit.

3. DNA S TRUCTURE & F UNCTION The manipulation of genetic information, DNA and RNA codes, is at the center of most biotechnology research and development.

4. The Central Dogma of Biology. Proteins are produced when genes on a DNA molecule are transcribed into mRNA, and mRNA is translated into the protein code. This is called “gene expression.” At any given moment, only a relatively small amount of DNA in a cell is being expressed.

5. DNA Structure. The nucleotides in one chain of the helix face one direction, while those in the other strand face the other direction. Each nucleotide contains a sugar molecule, a phosphate group, and a nitrogenous base. Nitrogenous bases from each strand bond to each other in the center through H-bonds. The H-bonds are rather weak; therefore, the two strands of DNA separate easily in high temperatures. DNA S TRUCTURE

6. N UCLEOTIDE S TRUCTURE DNA is composed of 4 nucleotides

7. S IMILARITIES IN DNA M OLECULES A MONG O RGANISMS 1. Virtually all DNA molecules form a double helix 2. The amount of adenosine equals the amount of thymine 3. The amount of guanosine equals the amount of cytosine 4. Nucleotides in each strand are oriented in the opposite direction of the other strand 5. Nitrogenous bases 6. DNA undergoes semi-conservative replication

8. DNA R EPLICATION DNA replicates in a semi-conservative fashion in which one strand unzips and each side is copied. It is considered semi- conservative since one copy of each parent strand is conserved in the next generation of DNA molecules.

9. V ARIATIONS IN DNA M OLECULES DNA varies in:  The number of DNA strands in the cells of an organism  The length in the base pairs of the DNA strands  The number and type of genes and noncoding regions  The shape of the DNA strands

10. S OURCES OF DNA In nature, DNA is made in cells. Mammalian Cell Culture Growing mammalian cells in culture is more challenging than growing bacterial cells Mammalian cells are grown in a broth culture Viral DNA Viruses are classified according to the type of cell they attack: Bacterial (bacteriophages) Plant Animal

11. P ROKARYOTIC DNA Bacterial Operon. An operon contains the controlling elements that turn genetic expression ON and OFF.

12. B ACTERIAL C ELL C ULTURE

13. E UKARYOTIC DNA Eukaryotic Gene. Eukaryotic genes have a promoter to which RNA polymerase binds, but they do not have an operator region. Transcription factors may bind at enhancer regions and increase gene expression.

14. I SOLATING AND M ANIPULATING DNA 1. Identification of the molecule(s) 2. Isolation of the instructions (DNA sequence/genes) for the production of the molecule(s) 3. Manipulation of the DNA instructions 4. Harvesting of the molecule or product, testing it, and marketing it

15. U SING G EL E LECTROPHORESIS TO S TUDY G ENE M OLECULES Gel Electrophoresis: Most commonly used when separating pieces of DNA no smaller than 50 bp and no larger than 25,000 bp The gel is “run” until molecules of different sizes are thought to have completely separated. Components of Gel Electrophoresis Powdered agarose Boiling buffer solution Running buffer DNA stain Sample load buffer

16. Agarose Gel Tray. Gel trays differ depending on the manufacturer. Each has some method of sealing the ends so that liquid agarose can mold into a gel. Some gel trays, such as those made by Owl Separation Systems, make a seal with the box, so casting a gel is simple. Other trays require masking tape on the ends to make a mold. Still others, like the one shown here, have gates that screw into position: up for pouring the gel and down for running the gel.

18. DNA A GAROSE G EL E LECTROPHORESIS

20. For the gel box to conduct electricity and establish an electric field with a positive end (red wire) and a negative end (black wire), the solution in the gel box must contain ions. The smallest molecules run fastest thru the gel

23. Q UESTIONS AND C OMMENTS ?

24. R EVIEW Q UESTIONS Your Turn! Put your name at the top of a sheet of paper, answer these questions and hand in: 1. Describe the relationship between genes, mRNA, and proteins. 2. Name the four nitrogen-containing bases found in DNA molecules and identify how they create a base pair. 3. The strands on a DNA molecule are said to be “antiparallel.” What does antiparallel mean? 4. During cell division, DNA molecules are replicated in a semi- conservative manner. What happens to the original DNA molecule during semic-onservative replication?

25. R EFERENCES 1. Biotechnology: Science for the New Millennium. 2012. Ellyn Daugherty.