Description / Overview of NGYX I.C. SmartSharp PCR DNA Ladder(s).
Let us assume that we would like to identify the optimal characteristics for a DNA Ladder covering the most usefull range (sizes) for PCR amplification products / restriction fragments analyses. Would you agree with:
Qualitatitvely speaking it should cover uniformly the range (sizes) starting with a 25-50 bp up to 5,0 Kb without any "gap" and with "easy to remember" + exact/precise fragments sizes.
It should deliver 2-3 quantitative scales also with "easy to remember" + exact/precise Molecular Weigth values.
Optimaly it should also give some possibilities to label it easily (i.e. RA, Fluorescence) or "probe it".
And last but not least, be very unexpansive to produce / easy to build up.
This may look somewhat a little bit ambitious... But NGYX I.C. did it for you! Here it is.
SmartSharp PCR DNA Ladder details.
Part I. Coverage and Quantitative Scales.
The picture on the right shows the SmartSharp PCR DNA Ladder (here referred as NGYX-PCR) together with world wide used "classic" DNA Ladders (Electrophoresis; EtBr staining).
Just have a look at it. Does it answer our first 2 requirements?
We have fragments from 25 bp (20 ng) up to 5,0 Kb (100ng). All sizes are exact/precise (design at bp level), easy to remember and uniformly distributed (Log-like overall scaling; no gap) over the full range.
Similarily we identify straight forward 3 Quantitative Scales (exact/presice as you will capture later from its production process) with also easy to remember MW values.
But wait the best still has to come...
Part II. Production and additional features.
The picture on the right shows both the SmartSharp PCR (NGYX-PCR) and the Smart DNA Ladders (Eurogentec; both at 50% load +/- 500 ng) in the same experimental conditions together with a single restriction fragment (XbaI) of the plamid pK7D4-DIY sized at 7.0 Kb.
Question: What's this pK7D4-DIY / XbaI? Why showing this fragment?
Answer: This fragment has been obtained from a plasmid pUC-Like "miniprep" (Qiagen) aliquot digested by XbaI Restriction Enzyme. And the plasmid name stands for p (pUC-like) K7 (7.0 Kb) D4 (4 Restriction Digestions) - DIY (Do-It-Yourself). This plasmid is also registered at BCCM under LMBP#4282.
And this plasmid is all what you need to build up the SmartSharp PCR DNA Ladders (we recommend to commercialize it EcoRI digestion + other digestions together; see picture below.
Good to know: this plasmid has antibiotic resistance gene (Spectino/Strepto) and a pUC amplification promoter. And it has been designed / created to incorporate about 100 restriction sites for 4 cheap/efficient, 5' protruding (labeling) Enzymes respectively: EcorI, HindIII, BamHI and XhoI. (plus XBaI if you want a 7.0 KB band)
Saying this you now capture for sure why the "DIY" stands for.
Part II. (continued).
Dubitative? Let us go for the following scenario.
Imagine that you are still graduating for i.e. your master in Molecular Biology. Your Lab's boss gives you a frozen sample (DH5 alpha Host Cells) of pK7D4-DIY and says: Can you produce the SmartSharp PCR DNA Ladder for our Lab needs (10 researchers) this year?
Sure! It is ONLY 48 hours culture (staying in control...2 Antibiotics Resistance), 1 working day to extract +/- 10 gr DNA (keep and save for several uses next years your e.g. Qiagen Columns) and another 1-2 working days to set-up RE digestions, clean these and put all together to have about 10,000 loads ready for use. Cost effective (all together: 0.10 Euro per Load)! Nice work. You just saved about 2,000 Euros for your lab this year! What if you would be a Producer / supplier working on larger scales: May be down to 0.05 Euro per load.
OK. We think that right now, reading these paragraphs there is not that much to add to convince you about the NGYX-PCR SamrtSharp. So there is probably only one question left: Where/How do I get the SmartSharp PCR DNA Ladder?