2x YT medium
2xYT is a nutritionally rich liquid microbial growth medium used for propagation of recombinant strains of Escherichia coli and M13 bacteriophage or other filamentous single-stranded DNA bacteriophages.
This medium contains a mainly enzymatic digest of casein and yeast extract.
Casein enzymic hydrolysate acts as a source of nitrogenous compounds like amino acids and peptides. Yeast extract supplies B-complex vitamins and minerals. Sodium Chloride maintains the isotonic condition of the medium. These components provide the necessary growth factors for rapid growth and replication of bacteriophage (without weakening the host cells) and Escherichia coli (allowing the bacteria to recover from the stress of transformation).
The pH of 2x YT medium is about pH 6.8 ± 0.2 at 25°C. pH can be adjusted using NaOH and HCl.
|Ingredients||For 100mL||For 500mL||For 1000ml|
|Tryptone||1.6 gm||8 gm||16 gm|
|Yeast extract||1 gm||5 gm||10 gm|
|NaCl||0.5 gm||2.5gm||5 gm|
|Distilled water||Upto 100 ml||Upto 500ml||Upto 1000ml|
- Weigh the ingredients separately with respect to the volume of the media. (Here, we are considering 1L of the media).
- Suspend the ingredients in a glass beaker containing about 1000mL of distilled water.
- Dissolve the components in the beaker using a magnetic stirrer. (Heat may be applied to dissolve the medium completely).
- Adjust the pH of the medium to the desired value.
- Transfer the broth to a conical flask or aliquot into smaller volumes.
- Close the mouth of the flask with a cotton plug. Seal it further with paper and a rubber band.
- Autoclave for 20 min at 15 psi (1.05kg/cm2) on liquid cycle.
- Mix well and pour into sterile Petri plates.
- Allow the medium to cool before adding, such as antibiotics (if needed).
Alternatively, the commercially available 2x YT agar media powders can be used. Weigh the mixture of content as prescribed by the manufacturer.
Performance: The media should show luxurious growth of Escherichia coli ATCC 23724 and Escherichia coli ATCC 53868 when incubated at 35-37°C for 18-24 hours.
Prepared Broth Media: Store at 2-8°C. Refrigerate if required, do not allow the products to freeze. Shelf life up to 6 months.
Many supplements, like antibiotics to the medium, are heat-sensitive, hence not autoclaved; instead, they are filter-sterilized and then added to the medium accordingly