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DNA转化
Chemical Transformation
· (Goldberg Lab)
Very nice protocol for E. Coli transformation including competent cell preparation and recipes
· (Crawford Lab)
· (Promega)
You can find general and useful protocol for the whole experiment of plasmid cloning from DNA and vector preparation, ligation, transformation, plasimd preparation and transfection...
· (Life Technologies)
How to stabilize unstable inserts and increasing plasmid yields
· (Life Technologies)
· (Chun-Ming Liu)
Quick E. Coli transformation method. It works!
· (Lazo Lab)
One-step PEG method
· Protocol for Transforming Competent E. coli Cells with Plasmid DNA (Dr. Chastain)
Very basic method for students.
- (Molecular Genetics Network Lab)
- Transformation of Aspergillus nidulans (Molecular Genetics Network Lab)
- (YGAC)
This protocol merges a typical LiAc protocol with the reducing agent component of the One-step method. It gives >103/ ug efficiency for our strain. Recipe is for 800 transformations (90 mls cells).
Electroporation
· Transformation of E. coli by Electroporation (Julie B. Wolf, UMBC)
provides details for preparing cell and DNA and electroporation procedure
· (Hoshi Lab)
· (Goldberg Lab)
Detailed protocol for preparing electroporation cell and electroporation procedure.
· Transformation of E. Coli by Electroporation (Goldberg Lab)
Very nice protocol for electroporation including cell preparation and recipes
· (Crawford Lab)
· (Life Technologies)
Preparation of Competent Cell
- (Donis Keller Lab)
Detailed method for competent cell preparation
- ) (NWFSC)
· ) (NWFSC)
· (PDF)(Templenton Lab)
A great method for making highly competent cells, simply.
· Preparation of Competent E. coli Cells (Waters Lab)
· (Crawford Lab)
· One-step competent E. coli cells (Dept. of Clinical Pharmacology, University of Berne, Switzerland)
A quick method of preparing competent cell. Transformation efficiency by this protocol is 106 to 107 cfu/mg.
· (PMCI Research)
Detailed protocol for cell preparation and electroporation
· (Hoshi Lab)
· (Bowtell Lab)
Including procedures for cell preparation and electroporation
Clone Screening and others
· (Donis Keller Lab)
To transfer DNA from bacterial colonies to nylon (or nitrocellulose) membrane so that a library can be screened by hybridization.
· (Herskowitz Lab)
Add colony directly into PCR tubes without plasmid preparation
- (Karl Clark)
Allows quick identification of correct clones if primers exist that allow clone identification by size or hybridization.
· Colony lifts
Lift plasmid colonies onto nitrocellulose filter for screen
· (Donis Keller Lab)
The toothpick assay is used to identify bacterial colonies containing recombinant plasmids following transformation and to obtain an approximation of the plasmid insert sizes.
· (Life Technologies)
For long-term storage of transformed cell
· (LTI)
Lawn cells require the F' episome for M13 infection and may be prepared using...
· (Crawford Lab)
Quick screening of transformants without minipreps
· (LTI)
· (Gerard R. Lazo)
Preserve E. coli strains in stab agar or glycerol for mid-term storage and lyophilize for long-term storage.
· (NUNC)
Positive clones are selected after plasmid cloning. This is often done by DNA probe hybridization in situ to plasmid DNA containing E.coli cells. The following protocol can be used for most plasmid cloning events in E.coli cells.
· (Hancock Lab) (Accessible only by IE)
This is a quick method to determine the presence of cloned fragments in plasmids after ligation/transformation.
· Media and buffers for Aspergillus transformation (Molecular Genetics Network Lab)
Transformation Trouble Shooting
· (LTI)
Very useful trouble shooting guide for common transformation problems like no colonies, all white or blue colonies, restriction resistance of your plasmid and more. If you are new to transformation, it's better to have a look.
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