Dehydrated Culture Medium
Enriched yeast extract–tryptone agar for high-density growth of recombinant Escherichia coli and M13 phage work
Cat. No. AS-139546 g/LpH 7.0 ± 0.2Non-selective agarMolecular biology
At a glance
- Target organismsE. coli host strains
- Incubation35–37 °C, aerobic
- Time16–24 h
- Sterilisation121 °C, 15 min
Manufactured in Melbourne. TDS and SDS available. Batch COA on request.
Overview
2× YT Agar (2× Yeast Extract Tryptone Agar) is an enriched, non-selective medium for growing recombinant Escherichia coli and for M13 bacteriophage work. The name comes from the original YT medium: 2× YT contains twice its tryptone and yeast extract (YT: 8 g/L tryptone, 5 g/L yeast extract).
Compared with LB Agar (Miller), 2× YT Agar has more tryptone (16 vs 10 g/L), twice the yeast extract (10 vs 5 g/L) and half the sodium chloride (5 vs 10 g/L). The richer nutrient base supports denser growth of E. coli host strains.
Key features
- Richer than LB: 16 g/L tryptone and 10 g/L yeast extract
- A standard medium for M13 phage work and phage display hosts such as XL1-Blue and TG1
- Accepts antibiotics and other heat-labile supplements added after autoclaving
Principle of the Medium
| Component | Function |
|---|---|
| Tryptone | Peptides, amino acids and nitrogen |
| Yeast extract | B vitamins, nucleotides and growth factors |
| Sodium chloride | Maintains osmotic balance |
| Agar | Solidifying agent |
The medium contains no selective agents. To select plasmid-bearing cells, add antibiotics such as ampicillin or kanamycin after autoclaving, once the medium has cooled to 45–50 °C.
Typical Composition
| Ingredient | g/L |
|---|---|
| Tryptone | 16.00 |
| Yeast extract | 10.00 |
| Sodium chloride | 5.00 |
| Agar | 15.00 |
| Total | 46.00 |
Final pH 7.0 ± 0.2 at 25 °C. Formula may be adjusted and/or supplemented to meet performance specifications.
Applications
| Field | Typical use |
|---|---|
| Molecular cloning | Growth and selection of transformed E. coli on antibiotic plates |
| Phage work | Propagation and titration of M13 bacteriophage; phage display with F′ host strains |
| Protein expression | Streaking and maintaining expression strains such as BL21(DE3) |
| Teaching laboratories | Transformation and plasmid selection practicals |
Comparable Products
| Manufacturer | Product | Relationship |
|---|---|---|
| HiMedia Laboratories | G035 2XYT Growth Agar | Same published formulation |
Frequently Asked Questions
What is the difference between 2× YT Agar and LB Agar?
2× YT contains 16 g/L tryptone and 10 g/L yeast extract, compared with 10 g/L and 5 g/L in LB (Miller). It also has half the salt (5 vs 10 g/L). The richer formula supports denser growth of E. coli.
Why is it called 2× YT?
It contains twice the tryptone and yeast extract of the original YT medium (8 g/L tryptone, 5 g/L yeast extract).
How much powder do I need?
46 g per litre of purified water. A 500 g pack makes about 10.8 litres, or roughly 430–540 plates of 20–25 mL.
When should I add antibiotics?
After autoclaving, once the agar has cooled to 45–50 °C. Adding them to hotter agar reduces their activity.
Intended use: For laboratory microbiological testing, quality control and research use only. Not intended for direct use in food, human or veterinary therapeutic applications, diagnosis, or consumption.
Mode of Action
Tryptone supplies peptides and amino acids, and yeast extract supplies B vitamins, nucleotides and other growth factors. The higher nutrient content than LB supports denser growth of E. coli. Sodium chloride maintains osmotic balance and agar solidifies the medium. The medium is non-selective; selection comes from antibiotics added after autoclaving.
Preparation
- Suspend 46 g of powder in 1 litre of purified or distilled water.
- Heat to boiling with frequent mixing until completely dissolved.
- Sterilise by autoclaving at 121 °C for 15 minutes.
- Cool to 45–50 °C in a water bath.
- Add filter-sterilised antibiotics or other supplements aseptically, if required, and mix gently.
- Pour 20–25 mL into each 90 mm Petri dish and allow to set on a level surface.
Liquid medium: 2× YT Broth uses the same formula without agar (31 g/L).
Incubation and Interpretation
| Parameter | Condition |
|---|---|
| Atmosphere | Aerobic |
| Temperature | 35–37 °C |
| Time | 16–24 hours |
| Positive result | Colonies on the agar surface |
Growth does not identify an organism or confirm a clone. Confirm transformants by colony PCR, restriction digestion or sequencing as appropriate.
Limitations
- Non-selective: contaminating organisms will also grow unless antibiotics are added.
- Antibiotics are heat-labile; adding them above 50 °C reduces their activity.
- Ampicillin plates lose activity during storage, and long incubation (over 16–20 hours) can produce satellite colonies.
- Growth rate varies between host strains.
Literature and References
- Sambrook J, Russell DW. Molecular Cloning: A Laboratory Manual, 3rd ed. Cold Spring Harbor Laboratory Press; 2001.
- Green MR, Sambrook J. Molecular Cloning: A Laboratory Manual, 4th ed. Cold Spring Harbor Laboratory Press; 2012.
- Messing J. New M13 vectors for cloning. Methods Enzymol. 1983;101:20–78.
- Barbas CF III, Burton DR, Scott JK, Silverman GJ. Phage Display: A Laboratory Manual. Cold Spring Harbor Laboratory Press; 2001.
- Atlas RM. Handbook of Microbiological Media, 4th ed. CRC Press; 2010.
Hazard Identification
Not classified as hazardous according to the GHS (Rev. 7) as adopted in Australia under the model WHS Regulations. Not Dangerous Goods under the ADG Code.
Fine powder can form combustible dust concentrations in air. Dust may cause mild mechanical irritation of the eyes and respiratory tract. Contains casein-derived material: people with milk protein allergy should avoid exposure.
Composition
| Ingredient | CAS No. | % w/w |
|---|---|---|
| Tryptone (enzymatic digest of casein) | 91079-40-2 | 34.8 |
| Yeast extract | 8013-01-2 | 21.7 |
| Sodium chloride | 7647-14-5 | 10.9 |
| Agar | 9002-18-0 | 32.6 |
First Aid
| Route | Action |
|---|---|
| Inhalation | Move to fresh air. Seek medical advice if symptoms persist. |
| Skin | Wash with soap and water. |
| Eyes | Rinse with water for several minutes. Seek medical advice if irritation persists. |
| Ingestion | Rinse mouth. Do not induce vomiting. Seek medical advice if unwell. |
Poisons Information Centre (Australia, 24 h): 13 11 26
Handling, Spills and Disposal
- Avoid generating and breathing dust. Weigh in a ventilated area.
- Spills: sweep up or HEPA-vacuum without raising dust; wash the area with water.
- Unused product: dispose of as non-hazardous waste under local regulations.
- Inoculated medium: treat as potentially biohazardous and decontaminate (for example by autoclaving) before disposal.
Quality Control Specification
Each batch is tested for growth performance. Aerobic incubation at 35–37 °C for 18–24 hours.
| QC organism | Strain | Inoculum (CFU) | Expected result |
|---|---|---|---|
| Escherichia coli | ATCC 25922 | 50–100 (≤ 100) | Good to luxuriant |
| Escherichia coli | ATCC 23724 | 50–100 (≤ 100) | Good to luxuriant |
| Escherichia coli K-12 | ATCC 23716 | 50–100 (≤ 100) | Good to luxuriant |
Physical and Chemical Tests
| Test | Specification |
|---|---|
| Appearance, powder | Cream to light yellow, homogeneous, free-flowing |
| Appearance, prepared medium | Light yellow, clear to slightly opalescent firm gel |
| Gel strength | Firm, comparable with a 1.5% agar gel |
| pH, prepared medium (25 °C) | 6.80–7.20 |
The information on this page is provided in good faith as a guide to the properties and use of this product and does not constitute a warranty of fitness for a particular purpose. Users are responsible for confirming suitability for their application. Ausamics Pty Ltd · ABN 56 676 640 467 · 36/25 Trafalgar Road, Epping VIC 3076, Australia · support@ausamics.com