I should do PCR, i have 100ug/ul but i don´t undestand how obtain 50ng in 50uL. Please can you help me?
I suposse that if use genomic dna i must use 50 ng.
Large lilutions should be done as serial dilutions. You have 100000 ng/µl. A volume of about 5 µl would be nice to pipet to the PCR mix, so each µl should finally have a conc. of 10 ng/µl. To get that conc., you would need to dilute the sample by the factor of 10000. Doing this in one step is a really bad idea (mix 1 µl of the sample with 9999 µl water to get 10 ml...!). Instead, make several smaller dilutions. Example:
1. mix 1 µl sample with 9 µl water to get 10 µl D1 (-> 10000 ng/µl).
2. mix 1 µl D1 with 9 µl water to get 10 µl D2 (-> 1000 ng/µl).
3. mix 1 µl D2 with 9 µl water to get 10 µl D3 (-> 100 ng/µl).
4. mix 1 µl D3 with 9 µl water to get 10 µl D4 (-> 10 ng/µl).
If you want to add less than 5 µl, you may use
4b. mix 5 µl D3 with 5 µl water to get 10 µl D4b (-> 50 ng/µl).
So that you can add 1µl of D4b to the PCR.
It is advisable to make dilutions in ranges that can be done with the same pipet (that means usually up to 1:10). You may make higher dilutions in a single step (like 1:100), but that will be less precise.
Have you tried using C1V1=C2V2?
http://www.mathcentre.ac.uk/resources/uploaded/c1v1c2v2.pdf
First you should change ug/ul to ng/ul (which is ug/ul times 1000.
Then will give you:100000ng/ul
Do a dilution 1:1000,
then if you have 100ng in 1ul (after 1:1000)
50ng in x
You cross, so you should take 50/100 as the volume which will give you 50ng,...
Large lilutions should be done as serial dilutions. You have 100000 ng/µl. A volume of about 5 µl would be nice to pipet to the PCR mix, so each µl should finally have a conc. of 10 ng/µl. To get that conc., you would need to dilute the sample by the factor of 10000. Doing this in one step is a really bad idea (mix 1 µl of the sample with 9999 µl water to get 10 ml...!). Instead, make several smaller dilutions. Example:
1. mix 1 µl sample with 9 µl water to get 10 µl D1 (-> 10000 ng/µl).
2. mix 1 µl D1 with 9 µl water to get 10 µl D2 (-> 1000 ng/µl).
3. mix 1 µl D2 with 9 µl water to get 10 µl D3 (-> 100 ng/µl).
4. mix 1 µl D3 with 9 µl water to get 10 µl D4 (-> 10 ng/µl).
If you want to add less than 5 µl, you may use
4b. mix 5 µl D3 with 5 µl water to get 10 µl D4b (-> 50 ng/µl).
So that you can add 1µl of D4b to the PCR.
It is advisable to make dilutions in ranges that can be done with the same pipet (that means usually up to 1:10). You may make higher dilutions in a single step (like 1:100), but that will be less precise.
Excellent explanation Wilhelm. Serial dilution is always better than one time dilution.
you may use C1V1= C2V2 formula to get your required concentration for any stock,
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