Gel ELECTROPHORESIS ?PLEASE HELP? - DNA ISOLATION (Aug/10/2006 )
i made a 0.8% agarose gel (dissolved 0.8g agarose in 100ml 1X TBE buffer)
Loaded 5 microliters of the genmic DNA (+ 5 micrliters loading buffer) as well as 50, 100 and 150 ng/microliter standards (i.e cncentration series). A high mlecular marker (ex. 1Kb ladder) which has been quantified was als loaded. A psitive contrl is also provided.
Elecrtophoreses the gel -- stain the gel with Ethidium Bromide--- photgraph under UV -light..
Im a little confused about the results :my Ladder goes up to 10 000bp but my DNA fragment is bigger than 10 000bp is this beacuse it is genomic DNA?
also the concentration series- I ive control, 50ng/microl, 100ng/microl and 150ng/microl
if i compare my DNA to the concentration series i think the concentration is 50ng/microliter
but could you please explain exactly what a concentration series is and also is it the concentratin of the ladder..
Also the positive control definitly works - could you please explain to me what more abut the positive control
yep. it is big because it is genomic
concentration series - concept is to provide you with examples of different concentrations of DNA to which you compare your sample in order to assess its concentration (based on brightness, not size) ie. the brigthness most similar to yours will give you an indication of your concentration
ladder is to give you an indication of size - some companies provide ladders with one or more bands at a specific concentrations, but this depends on how well you mix the tube before you load it on your gel!