Cell Cultures

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

timeline

Monday—LectureTuesday—procedures quiz for all,

Perform Demo

Prep.

Tuesday Lb broth base and LB agar base Several 250ml jars Bring in extra hood 10% BleachLab 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