26
votes
What does it mean to "write an image and GIF into the DNA of bacteria"?
The image was not in the DNA as such, only as an abstract representation that could be converted into an image from knowledge of the code. Briefly, they encoded the image into DNA, using a couple of ...
- 14.3k
17
votes
Accepted
What does it mean to "write an image and GIF into the DNA of bacteria"?
Just to add what might have been missing in the beautiful answer by @iayork. I just want to give a more simple picture of the encoding done in the E. coli DNA.
First for the rigid strategy in which 4 ...
- 14.1k
13
votes
Accepted
Assembling small DNA parts using Golden Gate
Purely going off experience here having used golden gate assembly methods for 5+ years now, there is a definite lack of literature regarding small part assemblies. In my current lab, we use CIDAR ...
- 316
11
votes
Accepted
Why is the start of my coding sequence ATG and not TAC?
This image from the Kahn Academy article 'Overview of transcription' might help:
Essentially, the sense/coding strand of the DNA encodes the sequence that is transcribed. The RNA polymerase binds to ...
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11
votes
Can I use multiple bicistronic RBS sequences in a synthetic biological circuit?
That’s a great question and has lot of opportunities to explore. I am not sure anyone has followed up on this original BCD work systematically. We did try cloning these elements on a medium copy ...
- 111
10
votes
Does gene orientation relative to origin of replication matter on small plasmids?
I like this question, and I had a similar thought when reading that "Forward or Reverse Strand" post. I've since found a 2005 publication by Mirkin and Mirkin1, which investigates the ...
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9
votes
Accepted
How would you use readthrough as part of a genetic circuit design?
Readthrough has already been used to implement transcriptional NOR gates, wherein tandem input promoters express a repressor that represses an output promoter. (First time I recall here: 21150903, ...
- 206
8
votes
Accepted
How long would you expect a designed plasmid to remain in a population of E. coli after transformation?
There isn't a simple answer.
There are a lot of factors which can influence how stable a plasmid will be. I'll describe what the main factors are, but to work out exactly how stable your plasmid is, ...
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8
votes
Measuring luminescence in a fluorescence plate reader
I am René Inckemann and I am PhD student at the Max Planck Institute in Marburg, Germany. In my PhD I am using luminescence on a regular basis and I was also responsible to organize some of our ...
8
votes
Accepted
Measuring luminescence in a fluorescence plate reader
As the other answers here say, technically plate readers which are capable of fluorescent measurements can also make luminescent readings, but the sensitivity may be low (https://www.biotek.com/...
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8
votes
Accepted
Why is electroporation not the prefered choice transforming for Mammalian Cells (HEK)?
Viral methods are just more efficient and give you a higher viability in the end. Electroporation works but you'll have a low yield of viable cells after. Here's a nice paper looking at the effects of ...
- 226
8
votes
Assembling small DNA parts using Golden Gate
I don't have hard data to share regarding you question, but I can provide some anecdotal evidence. I have used golden gate assembly to build hundreds of plasmids using ~20bp annealed oligos, as well ...
- 81
8
votes
Can we dilute PureExpress Cell Free Mix to increase number of reactions?
The PURExpress system from NEB is incredibly expensive, for this reason I have avoided it all costs during my experiments with cell-free systems.
This is the paper I've used to create my own cheaper ...
- 376
7
votes
Is there any problem with using a single stop codon with a single CDS in prokaryotes?
If there is only one CDS, presumably there will be a transcriptional termination directly after the CDS (after the stop codon actually), which makes any read-through coming from using a single stop ...
7
votes
Compatibility between spytag/spycatcher versions
I am currently working with these tags, so going off experience here. The binding efficiency is not 100% even when the correct versions are mixed e.g. version 2 spytag and version 2 spycatcher. I ...
- 316
7
votes
Accepted
Compatibility between spytag/spycatcher versions
I have worked with the coupling of SpyCatchers and SpyTags of different iterations and can confirm that they are all backward compatible. While SpyCatcher003 and SpyTag003 pairs have the quickest ...
- 86
6
votes
What does it mean to "write an image and GIF into the DNA of bacteria"?
Since a few people asked why the AAG triplet is avoided in the code, I thought I'd add this in addition to the other answers. The interesting part of this research ...
- 17.8k
6
votes
Accepted
Is there any problem with using a single stop codon with a single CDS in prokaryotes?
The impact of any read-through from a leaky stop codon in an expression unit with only one CDS would probably depend on a few things, mainly (i) where is the next in-frame stop codon, (ii) what are ...
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6
votes
Is there any problem with using a single stop codon with a single CDS in prokaryotes?
Perhaps I could add for eukaryotic systems which terminate all stop codons by a single protein, eRF1, this study by Schmied et al., showed readthrough of all stop codons to be less than 0.2% by wild-...
- 441
6
votes
Accepted
How can I convert plate reader measurements of Pichia pastoris OD to cells per ml?
From a technical perspective, for the FlopR package you mention (and I believe also for Jacob Beal's protocol) you can simply change the bead size and perform a new calibration. The only information ...
- 474
6
votes
Is there a favoured mathematical model of bacterial growth?
Some clarifications:
Commonly used models for bacterial isothermal growth curves (which represent the growth curve directly) are for example:
Exponential
Logistic
Generalized logistic
The Monod ...
5
votes
"Antibiotic resistance" equivalent in archaea for selection during cloning
Obviously, one can't use antibiotic resistance since they are not bacteria.
This is not so obvious. Some common antibiotics are active against archaea.1 The Halohandbook, an indispensable document ...
- 8,326
5
votes
Storage of bacteria
Aside from the comment that frozen cells require less manual maintenance and expense (which is a big plus), active cultures can rearrange/recombine plasmids causing mutations and deletions (especially ...
- 17.8k
5
votes
Is there a favoured mathematical model of bacterial growth?
the more biologically grounded Monod growth
Why is the Monod model more biologically grounded? It is empiric and NOT based on biological considerations! I only know this equation from enzyme kinetics,...
- 1,597
5
votes
Assembling small DNA parts using Golden Gate
It may also be worth considering how you are amplifying and isolating your small part. Gel extraction kits sometimes have an optimum DNA size ranges that may prevent recovery. Or if your part is ...
- 441
5
votes
Production of plant derivatives using genetic engineered micro-organisms
In principle, any biological product should be able to be developed through microbial synthesis, with the appropriate choice of chassis. Indeed, this was the goal of the DARPA "1000 Molecules&...
- 6,997
5
votes
Accepted
Which genetic oscillator should I use to generate oscillations in range of 2-20 mins?
I believe that what you are aiming to do is not possible with the mechanisms that you propose.
In most transcriptional and translational genetic circuits, the limiting factor for switching state is ...
- 6,997
5
votes
Accepted
DNA Design for Multi-Site Restriction Enzymes
In its active form, SacII (also called Cfr42I) is a homotetramer that must bind two copies of its recognition sequence in order to efficiently hydrolyze its substrate.1 Upon binding two sites on a ...
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4
votes
Storage of bacteria
Although a bacterial culture on a petri dish, slope or stab culture is in a closed environment and may start healthy, over time, the number of viable cells will decrease to zero as nutrients are being ...
- 1,519
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