Mapping Plasmids Circular 5Kb Plasmid?

Mapping plasmids?? Circular 5kb plasmid?

Restriction Enzyme Cuts:

EcoR1 1.0 1.7 2.3 BamH1 0.8 4.2 HinDIII 5.0 EcoR1 + BamH1 0.3 0.8 1.0 1.2 1.7 EcoR1 +HinDIII 0.5 1.0 1.2 2.3 BamH1 +HinDIII 0.8 1.8 2.4

The easiest way to visualize the mapping data is to draw three circles and then map each of the single digests to one of the three circles. Next take the simplest of the double digests and map this (BamHI + HindIII) and finally map the more difficult double digests and note that you may have two possible orientations of fragments (as you will see below). The simplest of the double digests shows you that you have two BamHI sites 0.8 kb from each other and the HindIII site is on the large 4.2 BamHI fragment which HindIII cuts off-center into two fragments of 1.8 and 2.4. Next look at the EcoRI + HindIII digest and you can see that the 1 kb and 2.3 kb fragments from the EcoRI single digest are unaffected, such that the 1.7 kb fragment is cut into two fragments (0.5 and 1.2) by HindIII.

So, your map so far (can't draw a circle, so we'll make this linear) is:

Single digests (in brackets is the same site as at the beginning)

HindIII --- 5 --- [HindIII]

EcoRI --- 1.0 --- EcoRI --- 1.7 --- EcoRI --- 2.3 --- [EcoRI]

BamHI --- 0.8 --- BamHI --- 4.2 --- [BamHI] Double digests

BamHI --- 0.8 --- BamHI ---- 1.8 --- HindIII --- 2.4 --- [BamHI]

EcoRI --- 1.0 --- EcoRI --- 1.2 --- HindIII --- 0.5 --- EcoRI --- 2.3 --- [EcoRI] OR EcoRI --- 1.0 --- EcoRI --- 0.5 --- HindIII --- 1.2 --- EcoRI --- 2.3 --- [EcoRI]

Finally, with the EcoRI and BamHI double digest, you keep the 1.0 and 1.7 fragments from the EcoRI single digest and the 0.8 kb fragment from the BamHI single digest. This means that both of the BamHI sites must be in the 2.3 kb EcoRI fragment, one is 0.3 kb from an EcoRI site and the other is 1.2 kb from the other EcoRI site flanking the 2.3 kb fragment:

EcoRI --- 1.0 --- EcoRI --- 1.7 --- EcoRI --- 0.3 --- BamHI --- 0.8 --- BamHI --- 1.2 --- [EcoRI] OR EcoRI --- 1.0 --- EcoRI --- 1.7 --- EcoRI --- 1.2 --- BamHI --- 0.8 --- BamHI --- 0.3 --- [EcoRI]

So, by choosing the second of the possibilities for the EcoRI + BamHI digest and placing the HindIII site properly by choosing the second of the possibilities of the EcoRI+HindIII double digest shown after the "OR" above, the overall map is:

EcoRI --- 1.0 --- EcoRI --- 0.5 --- HindIII --- 1.2 --- EcoRI --- 1.2 --- BamHI --- 0.8 --- BamHI --- 0.3 --- [EcoRI]