FACULTY OF ENGINEERING AND LIFE SCIENCES

UNIVERSITI SELANGOR

PRATICAL 1

COURSE : GENETIC ENGINEERING

COURSE CODE : BBS 2234
SEMESTER : APRIL 2023
LECTURER : MADAM YASOTHA SUNDARAJ
PROGRAMME : BACHELOR OF BIOTECHNOLOGY INDUSTRY

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PRATAYINI A/P PACKIANATHAN 4221003641 Pratayini
 PRATICAL 1

Preparation of Luria Broth Media Solution

Objective

1) The objective of this procedure is to outline the steps used to prepare Luria Broth
(LB) media to be used for bacterial culture growth in solution and bacterial colony
growth on petri plates.

Introduction

Luria Broth (LB) is nutrient-rich medium which is frequently used in laboratories to cultivate
microorganisms. To isolate individual (clonal) colonies of bacteria harboring a particular
plasmid, LB agar plates are usually utilized.The medium is employed for the general
cultivation and development of bacteria. Escherichia coli (E. coli) is frequently grown in it
for use in molecular biology applications and it supports the development of a wide variety of
bacteria.

Despite its widespread use today, Luria Broth Media Solution has its roots in the study of
bacteriophage genetics.The LB recipe was developed by Guiseppi Bertani in an effort to
maximise plaque development on a Shigella indicator strain. LB medium is a rich medium
that is frequently used for coliphage plaque assays as well as the cultivation of
Enterobacteriaceae members. In recombinant DNA research and other molecular biology
operations, LB and similar media are frequently employed. To select for cells that carry a
particular genetic component, such as a plasmid, a transposon, or a gene disruption via an
antibiotic resistance cassette, an antibiotic is frequently added to the sterilised medium.

Overview

a. A LB media solution is prepared using the following dry components:

i. Yeast extract
ii. Tryptone
iii. Sodium chloride (NaCl).

Procedure

A) Luria Broth Media

1. Each dry component of the solution were weighted out as follows:

i. Yeast extract: 5 g
ii. Tryptone: 10 g
iii. Sodium chloride (NaCl): 10 g

2. Weighed 5g of Luria Broth powder

3. Added each component to a 1 L Schott Bottle contained 200 mL of water

4. Dissolve components were stirred into solution.

 5. The solutions were autoclaved the solution at 121 °C to ensure that the LB is
sterilized of all foreign matter and contaminants.

6. The solutions were stored the solution in the refrigerator.

Apparatus and chemical

 Luria Broth powder

 Water
 Measuring cylinder
 Weighing scale
 Schott Bottle
 Magnetic Stirrer
 Magnet stirrer
 Autoclave

 LB Broth
 Nutrient Broth and Agar

Results

 Before Autoclave
  After Autoclave

Discussion

Due to its high nutritional value and straightforward composition, which can be easily
changed to meet specific needs, luria broth is typically utilised for molecular and genetic
studies. The modified version of Luria's original recipe is known as Luria Broth, according to
Lennox. Glucose addition aids in the preparation of Lennox's complete medium. Miller
reports that the sodium chloride content of Luria Broth is half that of Luria Broth. As a result,
the concentration of sodium chloride can be changed as desired. Recombinant E. coli strains
that are poor in producing B vitamins that were initially produced from E. coli strain K12 are
grown and maintained using luria broth. These stains are carefully altered to produce an
auxotrophic strain that cannot flourish on a nutritionally deficient medium.

Conclusions

As conclusion, the objective of experiment the is archived. This procedure describes how to
make Luria Broth (LB) media for bacterial culture growth in solution. Lysogeny broth is the
name given to the medium that is used to grow and develop bacteria in general. It helps a
variety of bacteria grow, including Escherichia coli (E. coli), which is frequently grown in it
for use in molecular biology applications. Furthermore, I have learn how to prepare the Luria
Broth media.
References

1. Sezonov, G., Joseleau-Petit, D. and D’Ari, R. (2007). Escherichia coli Physiology in Luria-
Bertani Broth. Journal of Bacteriology, [online] 189(23), pp.8746–8749.
doi:10.1128/jb.01368-07