EZgeneTM yeast plasmid kit is designed for rapid and reliable isolation of high-quality plasmid DNA from yeast cultures. Utilizing the reversible nucleic acid-binding properties of our matrix, the plasmid DNA is bound to the matrix while proteins and other unwanted impurities are eliminated by wash buffer. Pure DNA is then eluted. Purified DNA can be directly used in downstream applications such as PCR, restriction digestion, and Southern Blot.
The Yeast Plasmid Mini Kit combines the power of spin column technology with the lyticase, glass beads and alkaline-SDS lysis of yeast cells to yield high quality plasmid DNA in less than 1 hour. The mini spin columns facilitate the binding, washing, and elution steps, thus enabling multiple samples to be processed simultaneously. The actual plasmid yields depend on copy numbers, yeast strain, and conditions of growth. Because of low copy numbers, the maximum yield from 5 mL yeast culture is around 1 µg.
This protocol has been successfully used to isolate autonomous plasmids from S. cerevisiae. As a modified alkaline lysis procedure, genomic DNA is virtually eliminated from the preparation. Note that all centrifugation steps should be carried out at room temperature.
Storage and Stability:
All EZgeneTM Yeast Plasmid Mini Kit components are guaranteed for at least 12 months from the date of purchase when stored as follows: Buffer YP I/RNase A mix at 4°C, Lyticase at -20°C, all other components at room temperature.
Capacity:
The binding capacity for the mini column is 40 µg of plasmid DNA
Kit Contents:
Catalog#
BMG-YD1271-00
BMG-YD1271-01
BMG-YD1271-02
Preps
4
50
250
ezBindTM DNA Mini Columns
4
50
250
Buffer YPI
1.5 mL
15 mL
70 mL
Buffer YPII
1.5 mL
15 mL
70 mL
Buffer YPIII
2 mL
20 mL
100 mL
Buffer SE
3.0 mL
30 mL
135 mL
Buffer KB
3.0 mL
28 mL
135 mL
DNA Wash Buffer
2 mL
12 mL
50 mL
Glass beads
250 mg
2.7 g
13 g
Lyticase (units)
1100 U
11,000 U
55,000 U
RNase A
10 µL
50 µL
210 µL
Elution Buffer
600 µL
10 mL
30 mL
Manual
1
1
1
Important:
Dilute DNA Wash Buffer with absolute ethanol as follows：
YD1271-00: Add 8 mL absolute ethanol
YD1271-01: Add 48 mL absolute ethanol
YD1271-02: Add 200 mL absolute ethanol per bottle
Prepare a lyticase stock solution with Buffer SE and aliquot into adequate portions. Store each aliquot at -20°C and thaw before use. Each sample will require 30 µL of this solution.
YD1271-00: Dissolve with 130 µL Buffer SE
YD1271-01: Dissolve with 1.6 mL Buffer SE
YD1271-02: Dissolve with 8 mL Buffer SE
Add vial of RNase A to bottle of YPI and store at 4°C
Materials required but not supplied:
Tabletop micro-centrifuge and nuclease-free 1.5 mL tubes.
Water bath set to 30°C.
Absolute ethanol (96%-100%).
Before Starting:
Briefly examine this booklet and become familiar with each step. Prepare all components and have the necessary materials ready before starting.
