Cell Cultures Lab Manual 4e, 4f & 4g --Demo

timeline Monday—Lecture Tuesday—procedures quiz for all, Perform Demo

Prep. Tuesday  Lb broth base and LB agar base  Several 250ml jars  Bring in extra hood  10% Bleach Lab Service  Plates out

Write-up for 4e, 4f and 4g  Procedures—make an instruction page on how to pour plates

Safety Precautions  wear gogles

Tutorials  13 media prep.  14 lb agar and lb broth  15 introduction to sterile technique  17 streaking for isolated colonies  18 streaking for broth culture

Vocabulary Agar – solid media used for growing bacteria, fungi, plant, or other cells Media preparation – the process of combining and sterilizing ingredients (salts, sugars, growth factors, pH indicators, etc.) of a particular medium Autoclave – an instrument that creates high temperature and high pressure to sterilize equipment and media Enhancer – a section of DNA that increases the expression of a gene Intron – the region on a gene that is transcribed into an mRNA molecule but not expressed in a protein Exon – the region of a gene that directly codes for a protein; it is the region of the gene that is expressed Transcription factors – molecules that work to either turn on or off the transcription eukaryotic genes Histones – nuclear proteins that bind to chromosomal DNA and condense it into highly packed coils Nonpathogenic – not known to cause disease Bacteriophages – viruses that infect bacteria Gene therapy – the process of treating a disease or disorder by replacing a dysfunctional gene with a functional one

equations  Mass 1/vol 1 = mass 2/ volume 2

Lab 4E tips  Clean and label glassware!  Media is hygroscopic  Slowly stir in small amounts  LB broth has: tryptone, yeast extract and NaCl  LB Broth 20g/1L—do the conversion for the amount you are told to make  Use LB Broth base and add agar 1.5% to make LB agar (Example: 20g/L broth and 15g/1L)  We will pour the plates next class  Keep lid loose  Bottle size needs to be twice the volume of recipe

Lab 4F tips  Sterilize with 10% bleach spray  Re-heat LB agar in a microwave  pour plates in sterile environment when LB agar has cooled, so you can touch the glass  Leave plates in the hood  Plates will be used for lab 4g

Lab 4G tips  Bacteria waste goes into sterilizer tray, so it can be sterilized  Clam method