messi19

yeah there was something going on so went through my results again starting from the log till the end and its that lane 3 which seems to be producing the odd result but ill leave that and move onto the restriction map. il give it a go and then ask if i have any problems.thank you though for all the help

lane 2 - EcoRI & PstI digested - 3162 , 630 Lane 3 - BamHI& PstI digested - 2511,794 Lane 4 - BamHI& EcoRI digested - 3981, 501 Lane 5 BamHIdigested - 3981 Lane 6 PstIdigested - 3981 Lane 7 EcoRI digested - 3981

the log pairs converted to the standard is the antilog. my 3 double digests are roughly aaround my single digest.single digest is 3981. double digests are 3792, 3305 and 4482. so i take it the size of the plasmid is one of those from the double digest but how do u work out which one

the value i would be looking at is the log value surely as these do add up to the same value. but i dont use these for the restriction mapping? i use the antilog for the base pairs and mapping. so u only add up the smaller bands of the double digests to get the fragment size. im just getting confused so much with this.

Correct me if wrong but for the restriction map the sizes have to add up too the same value for each of the enzymes. But I don't believe mine do unless im using the wrong values

yeah did the inverse of the log , which gives me the fragment size of the logs. i presume these are the base pairs. for the single digests i have 3981. but i dont know what i add onto this to get the size of the plasmid cos i have 3 double digests which have smaller values such as 630,794 and 501 ahh . i believe i have figured it out . there is a double digest that also contains 3981 + 501. this would give me an answer of 4482 the other double digests are 3162 and 2511

oh right. ok well ill do the graph and the points now and let u know how i get on.. thanks a lot! did the graph as well as the curve and the interpolated the points to get the log. question 3 like u said is the sum of the parts. but which fragment sizes am i adding up. i take it its not the log fragment size so it should be the ones based on the gel?

http://imageshack.us/photo/my-images/52/scan0002fm.jpg/ thats the pic of the gel and the distances

thanks for the help. however when i start the graph with both points starting from 0 then i cant make a curve as the points are all located in one small area. so i decdied to plot a graph with migration distance going from 50 -60mm and log of fragment size going from 3.5 to 3.64. This was much better and did give me a curve. the next problem came when i had to Use the standard curve to determine the size of your pBR322 fragment bands from all the different combinations of enzymes - in Lanes 2-4 – and confirm the size of pBR322 for the single enzyme digests Lanes 5-7. the first 3 lanes contain 2 bands and the bands migration points are 104, 109 and 118mm. and these just cant be plotted on the graph as i dont have enough space or enough points to make the curve from. any idea on where im going wrong. just to be sure the migration distance goes on the x axis with the log on the y axis? thanx

having a problem with a few questions regarding the analysis of the plasmid. i have got the electrophoresis image and from there i have worked out the distance migrated for each band. it is from question 2 that i am unsure of what to do and how the graph should turn out. i would really appreciate any help i get. 1) From the photograph determine the migration distances (in mm) of the molecular weight standard DNA bands and of the bands of pBR322 cut with the different enzymes and construct a table with this data. i have done this question. and got the folowing results Fragment size (bp) Migration distance (mm) log of fragment size 3500 56 3.54 3200 59 3.51 4000 55 3.6 4300 53 3.63 4300 53 3.63 4300 53 3.63 2) Construct a standard curve (graph) of migration distance versus log molecular weight (in base pairs - bp) for the molecular weight standard bands plotting log10 marker fragment size (bp) against distance of fragment migration. Use the standard curve to determine the size of your pBR322 fragment bands from all the different combinations of enzymes - in Lanes 2-4 – and confirm the size of pBR322 for the single enzyme digests Lanes 5-7. Present your data in tabular form appropriately labelled. 3) What is the size of pBR322? How did you work this out from your results? 4) Using the data you have generated. Draw a representation of the plasmid showing the relative positions of the three restriction endonuclease sites. thank you