Biotechnology Explorer™ Microbial Culturing Kit Catalog #166-5020EDU explorer.bio-rad.com Store components of this kit at room temperature. Duplication of any part of this document permitted for classroom use only. Please visit explorer.bio-rad.com to access our selection of language translations for Biotechnology Explorer™ kit curricula. For technical support, call your local Bio-Rad office or, in the U.S.
Table of Contents Page Introduction........................................................................................................................i Kit Inventory Checklist ......................................................................................................1 Safety Issues....................................................................................................................1 Preparation of Liquid Media ....................................................................
Introduction The microbial culturing kit provides the materials and methods for a wide variety of microbiology activities. The kit can be used to make both liquid and solid media suitable for the growth of small or large cultures of either environmental microbes or laboratory strains of E.coli The microbial culturing kit is also part of the Bio-Rad Cloning and Sequencing Explorer Series (catalog #166-5000EDU) and contains the materials used in the Bio-Rad Microbes and Health kit (catalog #166-5030EDU).
Kit Inventory Checklist Kit Components Number/kit Ampicillin 2 vials LB nutrient agar powder (to make 500 ml) 1 pouch Petri dishes, sterile, bag of 20 2 bags Culture tubes 60 mm, sterile, bag of 25 3 bags Inoculation loops, sterile, 10 µl, pack of 10 loops 6 pks E.
Preparation of Liquid Media Preparation of Ampicillin Ampicillin is an antibiotic used to selectively eliminate bacteria that have not been transformed with plasmids containing an ampicillin resistance gene. Ampicillin is shipped freeze dried in a small vial containing 30 mg ampicillin. With a sterile pipet, add 3 ml of sterile water directly to the vial to rehydrate the antibiotic to make a 10 mg/ml or 200x solution.
Preparation of Ampicillin Broth This protocol is used to prepare liquid LB broth with ampicillin which will select for the growth of bacteria transformed with plasmids containing an ampicillin resistance gene. The final concentration of ampicillin in LB-Ampicillin broth should be 50 µg/ml. To make 25 mls of LB broth containing 50 µg/ml ampicillin, add 125 µl of 10 mg/ml ampicillin stock. Ensure LB broth is cooled to room temperature before adding ampicillin, excessive heat will degrade the antibiotic.
Preparation of Solid Media Preparation of Ampicillin Ampicillin is an antibiotic used to selectively eliminate bacteria that have not been transformed with plasmids containing an ampicillin resistance gene. Ampicillin is shipped freeze dried in a small vial containing 30 mg ampicillin. With a sterile pipet, add 3 ml of sterile water directly to the vial to rehydrate the antibiotic to make a 10 mg/ml or 200x solution.
Preparation of LB-Ampicillin Agar This protocol is used to prepare solid LB-ampicillin media which will select for the growth of bacteria transformed with plasmids containing an ampicillin resistance gene. The final concentration of ampicillin in LB-Ampicillin agar should be 50 µg/ml. To make 500 ml of LB agar containing 50 µg/ml ampicillin, add 2.5 ml of 10 mg/ml ampicillin stock. Ensure LB agar is cooled to 50°C before adding ampicillin, excessive heat will degrade the ampicillin.
Streaking Plates with Bacteria Streaking is done to make single colonies from concentrated bacteria. Each colony grows from one bacterium and thus a colony is a “clone” or group of genetically identical individuals. A tiny drop of the original bacterial suspension contains millions or billions of individual bacteria and must be diluted multiple times to isolate single bacteria. Under favorable conditions E.
Glossary Agar — Agar is a jelly-like substance obtained from seaweed. It is made of linked sugars (a polysaccharide) and is used to make medium for growing bacteria. Ampicillin — Ampicillin is a penicillin-like bactericidal antibiotic that inhibits the synthesis of the peptidoglycan component of bacterial cells walls especially in gram-positive bacteria but also in some gram-negative bacteria such as E. coli.
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