Use this category for questions directly related to DNA manipulation (isolation, purification, sequencing, etc.) and questions regarding general PCR methodologies
Moderators: r.rosati, mchlbrmn
how do i calculate expected size of fragments. , and the results for ddeI as below
DdeI sites in noname
# MS Cut position Site with flanks
---- -- ---------------- ----------------------------------------
1 12/15 2 catccaacat C^TCA_G catgatgaaa
2 35/38 25 aagctcctca C^TGA_G gactctgttt
3 329/332 319 atgtattacc C^TGA_G gacaaatatc
4 347/350 337 aaatatcatt C^TGA_G gggctgcatg
5 377/380 367 aaatatcaat C^TGA_G gggcccatcc
how they calculate? total bases is 385 bp
- Posts: 58
- Joined: May 30 2010 11:18 am
I don't really know what you are asking.
The 12/15 number means the first cut is in the top strand after base 12, and the bottom strand is cut after base number 15. Does that answer the question?
I'm sure you can find a more convenient format for the results from this or a different program.
- Posts: 3945
- Joined: Dec 13 2005 1:03 pm
- Location: Boston, USA
I really do not understand the meaning of these numbers. Is the DNA linear or circular? To calculate the fragment size, just calculate the number of bp between the two DdeI sites.
Retired academic researcher. Mention of a specific product does not imply my endorsement of the product. No conflict of interest or guarantee to work on the advice given. Do as I say, not as I do. Not liable to the loss of your valuable samples.
- PI of Posters
- Posts: 6991
- Joined: Jan 11 2006 12:40 pm
- Location: Mauna Kea
Is the DNA linear or circular?
What is its (uncut) size?
It looks to cut 5x. In the 5' strand (from the limited data you have given the frag sizes would be:
23 bp (cuts at 12 and 35 bp = 35-12)
294 (cuts at 35 and 329 bp = 329-35)
18 bp (cuts at 329 and 347bp=347-329)
and 30bp (cuts at 347 and 377bp= 377-347)
There will also be one ore more fragments depending on whether this is a linear or circular DNA.
(i.e. if linear the "ends are cut off" - one will be 12 bp = the distance from the start of the 5 strand to the first cut site and the second will be the distance from the 377 site to the 3' end of the DNA)
If circular plasmid there will be one fragment of size = (total plasmid size minus all the fragment sizes removed)
(Not forgetting the 12bp still attatched to the DNA before the first cut site).
I suggest you draw it out on paper, mark on the 5' cut sites (first number in the ddle cut info i.e. 12, 35, 329, 347, 377)
and calculate the DNA pieces cut out.
I am guessing that what has condused you is the two "site numbers" for each cut site: as mchlbrmn has mentioned these refer to the site of cutting on the 5' and 3' strands (you only need to consider 5' or first number to calculate the fragment sizes).
What I have written may seem needlessly complicated but we lack sufficient info to answer you question clearly (hence all of the caveats above).
- PI of Posters
- Posts: 515
- Joined: May 17 2010 7:13 am
If you use the "Custom Digest" function and select which enzymes you will cut with, on the results page, just click on "Fragments" under "List" in the bottom-right of the screen.
For this to work properly, you must specify whether the sequence is linear or circular (On the first sequence-entry screen)
- Posts: 12
- Joined: Mar 03 2012 3:36 pm
Return to DNA and General PCR Methods
Who is online
Users browsing this forum: No registered users and 1 guest