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Cloning pipeline does not work properly (Read 8419 times)
Feb 5th, 2011 at 5:29am

Peacemaker   Offline
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Hello,
at first, I would like to thank you for your great work on the program. It already offers many good features.
One very important feature for molecular biologists the cloning pipeline. Unfortunately, there seems to be a bug which makes this tool nearly unusable (I was using ugene-1.9.0-win-x86-r5586 under windows 7). The Problem is, when you clone some dna fragments with fitting sticky ends virtually (like most molecular biologists do), the program does a mistake during virtual ligation. As a consequence, one of the restriction sites used for cloning is no longer there after ligation. To demonstrate this, I cutted the Vector CVU55762.gb virtually using EcoR I and XbaI (menu cloning-digest into fragments). This results in two fragments with a EcoRI and XbaI overhang each.
Now, I performed a virtual ligation (menu cloning-construct molecule; both fragments were added; "Annotate fragments in new molecule" and "make circular" were checked). In theory, a molecule identical to the original vector should be the result. But after religation there is no more EcoR I restriction site, and the newly generated vector is not identical to the original one. Please see also the attached file new_mol.gb which I generated by this cloning process.
It would be very nice If you could fix that bug.
Best regards
 

new_mol.gb (9 KB | )
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Reply #1 - Feb 5th, 2011 at 3:34pm

Konstantin Okonechnikov   Offline
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Thanks for report!
This will be fixed ASAP

UPDATE: fixed in 1.9.1
« Last Edit: Feb 7th, 2011 at 1:25pm by Konstantin Okonechnikov »  
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Reply #2 - Oct 16th, 2012 at 12:35am

Anthony   Offline
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I am experiencing what seems like the same problem in version 1.11.2 (Windows 7 64 bit version). After I ligate fragments together, Ugene sets the origin of the plasmid at the beginning of one of the fragments in the middle of the cut site that is missing (KpnI). Is there anything special that I have to do in order to keep the annotation? I attached the molecules that I used in the cloning.

Thanks in advance,

Anthony
 

new_mol1.gb (5 KB | )
coelRFP.gb (65 KB | )
pUC19_rc.gb (5 KB | )
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Reply #3 - Oct 16th, 2012 at 1:04am

Anthony   Offline
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I should note that this problem only occurs when I reopen the new molecule file after closing the project. Immediately after ligation, the KpnI cut site is marked.
 
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Reply #4 - Oct 16th, 2012 at 3:21am

Konstantin Okonechnikov   Offline
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hi Anthony,
the KpnI sequence is still there.
Open new_mol.gb, then acticate circular view and search for enzymes again. (see screenshot)

The problem is the following: when the document with new molecule is saved the information about whether it is linear or circular is not recorded. So, when you open the project again, molecule is interpreted as linear DNA by default.

I've created the issue in the bug tracker, it will be fixed in 1.11.3.
 
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