Cloning vector

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roniadam
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Cloning vector

Post by roniadam » Fri Apr 04, 2008 2:21 am

Hi everyone:

how many enzymes are required to open a cloning vector for insertion of the cloned gene? I am using PFastBac vector and I cut it by one enzyme only. Is this will allow the interested fragment to insert into the vector or I have to use two enzymes???

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canalon
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Post by canalon » Fri Apr 04, 2008 9:33 pm

1 enzyme is enough but the risk is religation of the vector without integration of the fragment. And of course you wil not be able to select the orientation of your insert, which can be a problem if you plan to express a protein.
Patrick

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camzie
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Post by camzie » Thu Dec 04, 2008 6:35 am

i have a question:

Why isn’t the plasmid that contains the strongest promoter always the best expression vectors?

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MrMistery
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Post by MrMistery » Wed Dec 10, 2008 3:57 am

you might want to turn the expression on and off using an inducer...
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