Technique / Molecular Biology / Cloning library construction and screening


DNA Cloning Protocol



DNA Cloning Protocol

DNA cloning protocols content:

Preparation of oligo solutions
PCR experiments
Digestion of insert DNA
Digestion and dephosphorylation of vector DNA
Ligation of DNA fragments with sticky ends
Ligation of DNA fragments with blunt ends
Preparation of chemically competent E. coli cells
Transformation of plasmid DNA to chemically competent E. coli cells

(EMBL Hamburg)

Last update 10-Sep-2005, Rating of 8 votes.

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By ramya on 10-Mar-2009
can any one suggest where i would have gone wrong...
i have absolutely no colonies on plate after ligation...
TA cloning had enough colonies... but subcloning has no colonies... sequential digestion of vector was done and later gel purified and similarly was the insert prepared... and kept for lig for an hour at 25c and then transformed.. please help me... thanx in advance
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By Ram on 12-Nov-2008
You can do Restriction digestion. A single colony inoculate in 2ml LB broth for overnight. After 'miniprep' please take 1ul of plasmid, add 1Unit of desired restriction enzymes incubate for 2hours, analyse the result on electrophoresis
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By nadia on 10-Aug-2008
Please any body can suggest method for screeninig transformed colony after cloning expet PCR and colony Blot
Thanks alot
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By Mia on 19-Apr-2008
Does anyone have a protocol of shotgun cloning of Lambda-DNA fragments??

It's my first big experiment and I need help!! :(((

thanks
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By SHEETAL DHUMAL on 27-Oct-2007
THANKS FOR PROVIDING VERY NEAT PROTOCOL IN EACH SECTION OF MOLECULAR BIOLOGY WITH ADDITIONAL FACTORS AND IMPORTANT TIPS


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By mahdi on 11-Nov-2006
please i need the protocol of cloning in tandem

(Salmonella or E coli )

thank you

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By Fethi on 29-Apr-2006
please i need the protocol of cloning in tandem

(Salmonella or E coli )

thank you
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By ruchita selot on 10-Sep-2005
thanx a lot for providing with some of the most generally ignored tips while exp..
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