Lysogeny broth
Encyclopedia
Nutrient
A nutrient is a chemical that an organism needs to live and grow or a substance used in an organism's metabolism which must be taken in from its environment. They are used to build and repair tissues, regulate body processes and are converted to and used as energy...
rich medium
Growth medium
A growth medium or culture medium is a liquid or gel designed to support the growth of microorganisms or cells, or small plants like the moss Physcomitrella patens.There are different types of media for growing different types of cells....
, is primarily used for the growth
Bacterial growth
250px|right|thumb|Growth is shown as L = log where numbers is the number of colony forming units per ml, versus T Bacterial growth is the division of one bacterium into two daughter cells in a process called binary fission. Providing no mutational event occurs the resulting daughter cells are...
of bacteria. The acronym has been incorrectly interpreted as Luria broth, Lennox broth, or Luria-Bertani medium; according to its creator Giuseppe Bertani, the abbreviation LB was actually intended to stand for lysogeny broth. The formula of the LB medium was published in 1951 in the first paper of Bertani on lysogeny
Lysogeny
Lysogeny, or the lysogenic cycle, is one of two methods of viral reproduction . Lysogeny is characterized by integration of the bacteriophage nucleic acid into the host bacterium's genome...
. In this article he described the modified single-burst experiment and the isolation of the phages P1, P2
Enterobacteria phage P2
-Introduction:Bacteriophage P2 is a temperate phage that infects E. coli. It is a tailed virus with a contractile sheath and is thus classified in the family Myoviridae and the order Caudovirales, tailed viruses. Phage P2 has a double stranded DNA genome packaged in an icosahedral capsid with a...
, and P3. He had developed the LB medium to optimize Shigella
Shigella
Shigella is a genus of Gram-negative, nonspore forming, non-motile, rod-shaped bacteria closely related to Escherichia coli and Salmonella. The causative agent of human shigellosis, Shigella causes disease in primates, but not in other mammals. It is only naturally found in humans and apes. During...
growth and plaque formation.
LB media formulations have been an industry standard for the cultivation of Escherichia coli
Escherichia coli (molecular biology)
Escherichia coli is a gammaproteobacterium commonly found in the lower intestine of warm-blooded organisms and the descendants of two isolates are used routinely in molecular biology as both a tool and a model organism.-Diversity of Escherichia coli:Escherichia coli is one of the most diverse...
as far back as the 1950s. These media have been widely used in molecular microbiology
Molecular microbiology
Molecular microbiology is the branch of microbiology devoted to the study of the molecular principles of the physiological processes involved in the life cycle of prokaryotic and eukaryotic microorganisms such as bacteria, viruses, unicellular algae, fungi, and protozoa...
applications for the preparation of plasmid
Plasmid
In microbiology and genetics, a plasmid is a DNA molecule that is separate from, and can replicate independently of, the chromosomal DNA. They are double-stranded and, in many cases, circular...
DNA
DNA
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...
and recombinant
Recombinant
Recombinant may refer to:* A recombinant organism - an organism that contains a different combination of alleles from either of its parents.* Recombinant DNA - a form of artificial DNA* Recombinant virus - a virus formed by recombining genetic material...
proteins. It continues to be one of the most common media used for maintaining and cultivating laboratory recombinant strains of Escherichia coli. For physiological studies however, the use of LB medium is to be discouraged.
There are several common formulations of LB. Although they are different, they generally share a somewhat similar composition of ingredients used to promote growth, including the following:
- Peptides and caseinCaseinCasein is the name for a family of related phosphoprotein proteins . These proteins are commonly found in mammalian milk, making up 80% of the proteins in cow milk and between 60% and 65% of the proteins in human milk....
peptonesPeptidePeptides are short polymers of amino acid monomers linked by peptide bonds. They are distinguished from proteins on the basis of size, typically containing less than 50 monomer units. The shortest peptides are dipeptides, consisting of two amino acids joined by a single peptide bond... - Vitamins (including B vitamins)
- Trace elements (e.g. nitrogen, sulfur, magnesium)
- Minerals
Peptides and peptones are provided by tryptone
Tryptone
Tryptone is the assortment of peptides formed by the digestion of casein by the protease trypsin.Tryptone is commonly used in microbiology to produce Lysogeny broth for the growth of E. coli and other microorganisms...
. Vitamins and certain trace elements are provided by yeast extract. Sodium ions for transport and osmotic balance are provided by sodium chloride
Sodium chloride
Sodium chloride, also known as salt, common salt, table salt or halite, is an inorganic compound with the formula NaCl. Sodium chloride is the salt most responsible for the salinity of the ocean and of the extracellular fluid of many multicellular organisms...
. Bacto-tryptone is used to provide essential amino acid
Amino acid
Amino acids are molecules containing an amine group, a carboxylic acid group and a side-chain that varies between different amino acids. The key elements of an amino acid are carbon, hydrogen, oxygen, and nitrogen...
s to the growing bacteria, while the bacto-yeast
Yeast
Yeasts are eukaryotic micro-organisms classified in the kingdom Fungi, with 1,500 species currently described estimated to be only 1% of all fungal species. Most reproduce asexually by mitosis, and many do so by an asymmetric division process called budding...
extract is used to provide a plethora of organic compound
Organic compound
An organic compound is any member of a large class of gaseous, liquid, or solid chemical compounds whose molecules contain carbon. For historical reasons discussed below, a few types of carbon-containing compounds such as carbides, carbonates, simple oxides of carbon, and cyanides, as well as the...
s helpful for bacterial growth.
In his original 1951 paper, Bertani used 10 grams of NaCl and 1 gram of glucose per 1 L of solution; Luria in his "L broth" of 1957 copied Bertani's original recipe exactly. Recipes published later have typically left out the glucose.
Formulas
The formulations generally differ in the amount of sodium chloride, thus providing selection of the appropriate osmotic conditions for the particular bacterial strain and desired culture conditions. The low salt formulations, Lennox and Luria, are ideal for cultures requiring salt-sensitive antibiotics.- LB-Miller (10 g/L NaCl)
- LB-Lennox (5 g/L NaCl)
- LB-Luria (0.5 g/L NaCl)
Preparation
The following is a common method for the preparation of 1 liter of LB:- Measure out the following:
- 10 g tryptoneTryptoneTryptone is the assortment of peptides formed by the digestion of casein by the protease trypsin.Tryptone is commonly used in microbiology to produce Lysogeny broth for the growth of E. coli and other microorganisms...
- 5 g yeast extract
- 10 g NaCl
- 10 g tryptone
- Suspend the solids in ~800 ml of distilled or deionized water.
- Add further distilled or deionized water, in a measuring cylinder to ensure accuracy, to make a total of 1 litre.
- AutoclaveAutoclaveAn autoclave is an instrument used to sterilize equipment and supplies by subjecting them to high pressure saturated steam at 121 °C for around 15–20 minutes depending on the size of the load and the contents. It was invented by Charles Chamberland in 1879, although a precursor known as the...
at 121 °C. - After cooling, swirl the flask to ensure mixing, and the LB is ready for use.
Adjusting the pH
Prior to autoclaving, some labs adjust the pH of LB to 7.5 or 8 with sodium hydroxide. However sodium hydroxide does not provide any buffering capacity to the media, and this results in rapid changes to the pH during bacteria cultivation. As a result, some labs adjust the pH of LB with 5–10 mmol/L TRISTris
Tris is an abbreviation of the organic compound known as trisaminomethane, with the formula 3CNH2. Tris is extensively used in biochemistry and molecular biology. In biochemistry, tris is widely used as a component of buffer solutions, such as in TAE and TBE buffer, especially for solutions of...
buffer, diluted from 1 mol/L TRIS stock.
High density bacteria cultures require more buffering capacity than 5-10 mM TRIS can provide.
See also
- Agar plateAgar plateAn agar plate is a Petri dish that contains a growth medium used to culture microorganisms or small plants like the moss Physcomitrella patens.Selective growth compounds may also be added to the media, such as antibiotics....
- Salvador Luria
- SOC mediumSOC mediumSuper Optimal Broth is a nutrient-rich bacterial growth medium used for microbiological culture, generally of Escherichia coli. It was developed by Douglas Hanahan in 1983 and is an adjusted version of the commonly used LB media . Growth of E...
- perhaps the second-most widely used medium for culture of Escherichia coliEscherichia coliEscherichia coli is a Gram-negative, rod-shaped bacterium that is commonly found in the lower intestine of warm-blooded organisms . Most E. coli strains are harmless, but some serotypes can cause serious food poisoning in humans, and are occasionally responsible for product recalls...
in molecular biologyMolecular biologyMolecular biology is the branch of biology that deals with the molecular basis of biological activity. This field overlaps with other areas of biology and chemistry, particularly genetics and biochemistry...
work