Ape plasmid editor manual
Jun 02, · Quick Guide to ApE (A plasmid Editor) Since I prefer to clone my riboprobes, rather than amplifying them every time from cDNA, I have to making in silico maps of the constructs first. To do so I use a freely available software called ‘A plasmid editor’ or ApE. Below is a description of how I go about making my plasmid maps. Nov 03, · A brief tutorial showing how to use APE for basic cloning experiments.
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Below is a description of how I go about making my plasmid maps. Once you have installed the program, this is the kind of window you will get.
There is a main toolbar, in my case at the top of the screen, and there are icons in the window, where the sequence is pasted. First I copied and pasted the sequence of the vector into the empty window of ApE. Since this is a plasmid, I set it to circular. The diagram tells us, that the insertion site is flanked by two EcoRI restriction sites.
These are the only two EcoRI sites in the plasmid. The numbers next to the name of the enzyme indicate, how many times the enzyme cuts in the construct.
If you press this button, the program will show, where in your sequence the restriction sites are for the enzymes you have selected.
You can select and de-select an enzyme simply by clicking on it. However, this might not be the ideal probe for in situ hybridization, since I have not included any of the UTRs or checked for alternate transcripts. Note: Do not forget to save your files individually.
If you make changes to your original vector, safe it as a separate file. Once I have saved the sequence of the insert in a separate file, I can copy it and past it at the correct site in my vector. Which then looks something like in the image below. When you click on it a drop down menu opens.
You can give a name to the feature you added blue arrow.
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It is also possible to specify the feature type green arrow. Is it a gene, a primer, a binding site or something else. It is also very useful to give your feature a unique color. Once you are done press OK. When you go to the graphic map of your plasmid, you get something like in the image below. The EcoRI restriction sites are still highlighted in the list and I also added the insertion site as a feature in the map. To account for that, I have to make two versions of the final construct.
Above I have shown you the forward construct. Once the sequence is converted, it has to be pasted in the vector as described above and saved as a separate file.
ApE- A Plasmid Editor
Now I can annotate the features and give it a color. But to determine the orientation of the insert, I have to look for a unique restriction site, which is towards one end of my insert and which is also present in the vector.
The reason will become apparent soon. As you can see in the graphic map of my insert which is linear by the way , I have picked DraI. It cuts towards one end of the fragment. It allows you to run a virtual gel. As you can see in the image below, depending on the orientation of my insert, the band pattern of the digest changes. For the forward orientation I am supposed to get a bp and a bp band.
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Personally, I prefer to use the T7 polymerase, so I am looking for inserts in forward orientation. Remember, you want the antisense probe.
Once you have confirmed that you cloned your insert and you determined the orientation, it is a good practice to send the construct for sequencing. But if you are impatient like me, you can straight away get down to preparing your probe and testing it out. By the way, this program is more powerful, than what I just described.
Below are the tools available in the current version. What version of this program do you use?
I can see that this program is very powerfull for develoment vectors of clonation. Thanks and good night. Like Like. I am using Version v2. Yes, indeed it is a fantastic and very powerful software. The best thing is, that it is free.
Quick Guide to ApE (A plasmid Editor)
How can I add multiple sequences in a window of APE for analysis. Regards, Shahbaz. Hi Shahbaz, I am not sure I understand the question. Do you want to blast two sequences?
Best Simoe. I am a first time user and this is great. Many Thanks for posting this this one. Like Liked by 1 person. Hi Chatham, thank you for your kind words. Is there anything in particular you would like to know about?
Best regards Simone.
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Notify me of new comments via email. Notify me of new posts via email. I hope you liked my brief description. Share this: Twitter Facebook. Like this: Like Loading Next Article cDNA synthesis.
Hi Edgar, I am using Version v2. Best regards Simone Like Like.
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