User:Jarle Pahr/Questions
Technical and scientific questions which can conceivable by answered by in silico or small-scale, low-cost wetlab experiments:
DNA purification/isolation:
How does additional washing steps affect the purity of miniprep DNA samples?
What is the difference and variation in yield for 6 mL cultures grown in 13 mL tubes and ErlenMeyer bottles, respectively?
How long must cultures be grown to achieve acceptable plasmid yields? Is 12 h enough?
Experiment: Inoculate several typical miniprep-scale cultures, incubate 12 h, perform miniprep, observe yields.
Ligation:
How does the probability of insert concatenation vary with insert concentration/molar amount (see Simulations)?
Transformation:
What is the smallest amount of plasmid DNA that reliably yield transformants?
What effect does snap-freezing have on the transformation efficiency of supercompetent cells?
Experiment: Make one batch with half snap-freezed, the other half not snap-freezed, determine transformation efficienies for both conditions.
What variable (pre heatshock-incubation, post heat shock-incubation) has the largest effect on transformation efficiency (and how large is the effect)?
How does CFU vary with post heat shock incubation time?
SLIC:
Can small (~100 bp fragments) be succesfully cloned by SLIC? How? Suggestions: Use klenow polymerase, or very short incubation times. (add dCTP after 0 seconds, 10 s, 30 s, etc.). Klenow fragment has both 3'-5' exonuclease and 5'-'3 polymerase activity, so stopping the reaction with dCTP should work same as with T4 DNA polymerase.
PCR:
How does PCR yield vary with annealing temperature for a given reaction?
Experiment: Set up a gradient PCR and observe band intensity for the various positions. Or run several subsequent reactions with different annealing temperatures.
Misc:
Non-standard growth media: Can E. coli grow on "dextro" energy tablets?
Verification of commercial personal-genomics results: Can DNA extraction, PCR and sequencing confirm a SNP result reported by 23andme?
Commercial kit performance: What are the performance properties of various commercial molecular-biology kits?
Can horse meat be reliably detected by a cheap, at-home PCR test?
Do dNTPs show up during gel electrophoresis?
pllD: Can activity of the pllD promoter be shown by expression of GFP in a minimal medium?
Given a certain mutation rate, for how long can one expect maintain 100 % sequence preservation in a plasmid.
When working with plasmids, a very large number of DNA molecules is handled simultaneously, but mutations occur in discrete molecules. What are the "population dynamics" of mutations?
What organisms live in a home environment? Can culturing, PCR and 16S sequencing be used to identify those organisms?
Does molten agarose contract or expand when solidifying?
LA medium darkens when stored in liquid form (warm) before pouring. What is the cause of this? Does it affect the nutritional properties of the medium?
Culturing and bacterial growth:
Does the initial cell density upon inoculation affect the final cell density?
How long does it take to reach maximal cell density for typical miniprep culturing ( ca. 3 mL in 13 mL tube, for example)
How long is a plasmid typically retained in the absence of selection pressure?
Big questions
Oligonucleotide synthesis: Will there be further radical improvement?
Is Cambrian Geonomics it?
Open questions in biology: http://www.biomedcentral.com/bmcbiol/series/openquestionsinbiology
