Team:UNITN-Trento/Notebook/Labposts/06/55
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{ "date" : "2013-06-25", "author" : "emil", "title" : "Purification and digestion of R0010", "content" : "I purified, and then quantified, 3 out of 5 of the previous inocula following the miniprep protocol with vacuum.
Quantification results
Afterwards, I digested 800ng of each sample with Spe1-HF and Pst1-HF following the digestion protocol for screening. Finally I ran the products on a gel (1,5% agarose, due to the short length of the insert: only 200 bp + prefix and suffix). I loaded 12µl of a ladder 100bp prepared as follows:Sample | Quantity |
---|---|
1 | 396.1ng/µl |
2 | 523.1ng/µl |
3 | 171ng/µl |
Ladder
This time, I loaded 12µl (10µl of sample and 2µl of glicerol solution 30%) of each sample.Quantity | |
---|---|
Water | 4µl |
Sharpmass Euroclone 100bp ladder | 1µl |
glicerol solution(30%) | 1µl |
Gel order
We can see 2 bands: the highest is the backbone (pSB1A2: 2079bp) and the lower is pLac (nearly 243bp, located between the bands 300 and 200bp of the ladder).Sample | Well |
---|---|
2µl loading dye | 1 |
Ladder | 2 |
sample 1 | 3 |
sample 2 | 4 |
sample 3 | 5 |
sample 4(Bruno e Fabio) | 6 |
sample 4(Bruno e Fabio) | 6 |
Ladder 1000 kb | 7 |
loading dye | 8 |
After the screening I digested 3µg of the pSB1A2+R0010 (sample #2) with SpeI and PstI-HF o/n following the usual protocol.", "tags" : "Plac" }