Plasmid Restriction Maps

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sims10
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Plasmid Restriction Maps

Post by sims10 » Mon Nov 27, 2006 12:01 am

I'm having problem drawing a plasmid map with the following combination enzymes...

BamH1: 900/1500/3000
Hindlll: 500/1600/3300
BamH1 + Hindlll: 300/400/500/1200/3000

Is there a simple way to solve this?

sdekivit
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Post by sdekivit » Mon Nov 27, 2006 8:28 am

first you gave to draw the restriction fragments from the separate enzymes. Thens you can take a look on where the restriction sites for both enzymes located from the combined enzyme data.

2810712
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Post by 2810712 » Mon Nov 27, 2006 9:26 am

yeah, the way above said get to the ans.
see the no. of frags = no. of sites
so first and second both have 3 sites each, but the combined case has not 3+3=6 but 5, so they sahre one site...

now the combined result can be treated as one obtained by two stepped restriction...i mean imagine 1st BamH1 gives u 3 frags...
now u treate these 3 frags with the Hindlll, so u get the above said 5 frags ,
so u need to cut those only-BamH1 frags into those 5 frags...
you can form only smaller or equal length frags from them, so u get this constiant,
here Bam1 gives us 3000,1500,900
and the combined treate gives 300,400,500,1200,3000

now the 3000 seems remain intact,
while 900 frag seems get broke into 300, and 400 frags
and the 1500 frag into the 1200 and 300 strands

now u get the possible distances between BamH1 and Hindlll sites...
now try to make out the only Hind result from this to narrow down the possible plasmid map possibilities...

or

from combined result try to make the 'only' results by associating diff. strands togather...this is faster,


to get Hindlll result
500 is intact, 400 and 1200 associate, 300+3000 associate
so the sequence is -[B/H]500[H] 400 [B] 1200 [H] 300[B] 3000-
[B]= BamH1 site
[H]= HIndlll site
[B/H]= both



it ws nice exercise, thanks...

sims10
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Post by sims10 » Mon Nov 27, 2006 5:09 pm

ok so I tried solving it...can you check if I'm correct...

so at the top is 5400 bp (BamH1 site)

starting counter-clockwise...

500 bp (Hindlll)
900 bp (BamH1)
2100 bp (Hindlll)
2400 bp (BamH1

2810712
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Post by 2810712 » Tue Nov 28, 2006 8:40 am

umm...
i think instead of that 900 term u mean 400, as otherwise the total length gets more than 5400...
try to recheck ur answer by predicting the fragment lengths on digestion using the plasim map u suspected...

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