The number 496 is a positive integer that is often known for its interesting property in mathematics: it is a *perfect number*. A perfect number is defined as a positive integer that is equal to the sum of its proper divisors, excluding itself. For 496, the proper divisors are 1, 2, 4, 8, 16, 31, 62, 124, and 248.

More generic PCR information at: Section "Polymerase chain reaction".

Because it is considered the less interesting step, and because it takes quite some time, this step was done by the event organizers between the two event days, so participants did not get to take many photos.

PCR protocols are very standard it seems, all that biologists need to know to reproduce is the time and temperature of each step.

We did 35 cycles of:

This process used a Marshal Scientific MJ Research PTC-200 Thermal Cycler:

We added PCR primers for regions that surround the 16S DNA. The primers are just bought from a vendor, and we used well known regions are called 27F and 1492R. Here is a paper that analyzes other choices: academic.oup.com/femsle/article/221/2/299/630719 (archive) "Evaluation of primers and PCR conditions for the analysis of 16S rRNA genes from a natural environment" by Yuichi Hongoh, Hiroe Yuzawa, Moriya Ohkuma, Toshiaki Kudo (2003)

One cool thing about the PCR is that we can also add a known barcode at the end of each primer as shown at Code 1. "PCR diagram".

This means that we bought a few different versions of our 27F/1492R primers, each with a different small DNA tag attached directly to them in addition to the matching sequence.

This way, we were able to:

Input: Bacterial DNA (a little bit)
... --- 27S --- 16S --- 1492R --- ...

|||
|||
vvv

Output: PCR output (a lot of)
Barcode --- 27S --- 16S --- 1492R

Finally, after purification, we used the Qiagen QIAquick PCR Purification Kit protocol to purify the generated from unwanted PCR byproducts.

It has to swim fast, right!

So how is it that mitochondria are not inherited from the father?

Ciro Santilli is actively looking for donations and contracts so he can continue to work full time on OurBigBook.com sustainably, and develop free hardcore university-level STEM education for all ages!

For 300k USD I will quit my job or not get a new job and work on OurBigBook full time for a second year to try and kickstart The Higher Education Revolution. Status: ~44k / 300k USD. At 2M USD I retire and work on open STEM forever.

I first quit my job 1st June 2024 to work on the project for 1 year after I reached my initial 100k goal mostly via a 1000 Monero donation. In this first year I improved project tech, but didn't go and solve university courses to create super targeted content, and didn't obtain a single contributing user except myself, see a summary at Section "OurBigBook Project Update March 2024".

For a followup, it would be an interesting experiment to spend on year full time solving as many courses as I can from a world class university in the city where I live in the UK, and adding that as content to the platform to see if that would attract interest, and reaching out directly to course takers at their university environment to try and help them. I intended to do that in year one but my got distracted by tech. It is quite possible that no one has ever done that before in history: a highly motivated technical person with the time and opportunity to do one single thing: help top university students learn their STEM courses better and have more fun doing so. I have in particular identified one course where this would be particularly feasible: the mathematics course, given that much of their course materials, and also their building are quite open. I do sometimes wonder if doing this would be just a waste of my life. But part of me tells me it could generate big interest and is worth a try.

For a second follow up year, I increased my requirement to 300k USD to give me more peace of mind. So the total donation so far is 244k, and if I reach a total of 300k USD, then I'll work on the project for a second year.

At 2M USD I retire and work on open STEM projects forever. At these timelines, I can't guarantee it will be specifically on education technology specifically all the way, but I guarantee that whatever it is it will be open and extremely well explained as usual.

Total donations to date ~244k USD. Donation breakdown:More details: Section "Accounting method"

How to give:And if you have a different preferred payment mechanism not listed above, please contact Ciro, and he will set it up.

Ciro's current ambitions require him to remain in developed countries, because Ciro wants to document advanced science and technology by liaising with top universities, and there is not nearly as much high technology in poor countries. Remaining in developed countries is also a required due to family reasons.

Note to potential anonymous crypto donors: I live in the UK, and after some messy back and forth that included freezing my account at one point, Barclays finally decided that they do not allow me to receive anonymous donations. And I'm pretty sure that the same would happen on any other British bank sooner or later. Therefore, while I continue to accept anonymous donations, they will not count towards my "if I get X I do Y"-type donation goals, since I can't reliably spend that money. If you want to donate in crypto, and clearly give me your real identity and an explanation of how you got the money, then that is fine, it can count. Just be warned that I will need to give that information to my bank and clear it with them beforehand, and if they are still not happy with it, I'll just give it all back. Your identity does not have to be publicly disclosed, only the dates and amounts. But my bank has to know.

If you would like public acknowledgement for your support, Ciro will very gladly give it, just let Ciro know how you'd prefer it. Due to Ciro Santilli's campaign for freedom of speech in China, many supporters have chosen to be anonymous, and that is totally fine, not everyone is interested in politics, or has a situation where going public is acceptable, so we don't have a standard setup yet, let's build it together. A acknowledgement section at the bottom of this page would be a minimum, but I for larger donations we could add a your advertisement in a locations such as:

100k USD/year is a semi arbitrary amount that sounds nice. My last day job total compensation as of 2024 was about 150k USD/year.

An ecotone is a transition area between two different ecological communities or ecosystems. These areas often have a mix of species from both adjacent communities, and they can exhibit unique ecological characteristics and higher biodiversity compared to the surrounding areas. Ecotones can occur at various scales, such as between forests and grasslands, wetlands and uplands, or even between different types of aquatic systems.

A protein that is a catalyst for some chemical reaction.

For an initial concrete example, consider e. Coli K-12 MG1655 gene thrA.

Mission: to live in a world where you can learn university-level mathematics, physics, chemistry, biology and engineering from perfect free books that anyone can write to get famous.

Live website: ourbigbook.com

Further information and rationale: Section "OurBigBook.com"

The project's mission is one of, or perhaps the most important, life objective of Ciro Santilli. Reproductive goals aside. These two types of goal are incommensurable. This is one of the great challenges of life.

This is ongoing project.

Ciro's goals in advertising this half done project are is partly to obtain some feedback, and partly to give the idea to someone else who might help push it further, be it in this stack or not.

Better editing support is a must, likely WYSIWYG.

But besides that, it is already in broad strokes the best approach Ciro Santilli can come up with to try and reach the mission statement only with technical advances, i.e. without large amounts of money or political influence which Ciro Santilli does not have.

Maybe that website isn't enough of a technical advance to reach its mission. Maybe there is some further not yet imagined technical insight that would push it into viability. Maybe not. But one must try. Only God can know the answer to these questions.

As of 2022, Ciro has spent about 2.5 years full time working on this project. First he spent about 1 year in 2014 on the first iteration: github.com/booktree/booktree, a GitLab fork, but then decided it was not the way to go.

Then around 2021 he put in some more 1.5 year of full time work, now with a possibly overly complicated (or perhaps just insane/immature) Next.js/Sequelize from scratch website stack.

It makes Ciro a bit ashamed to see that "so little user visible stuff was achieved in so much time". It is partly because he and many people underestimate the difficulty of web development. Perhaps there were some bad stack/useless feature choices issues. And a good dose of indulging in studying the natural sciences to bootstrap content and have fun. But really trying is the only way to learn.

See also: github.com/cirosantilli/awesome-whole-cell-simulation

As of 2019, the silicon industry is ending, and molecular biology technology is one of the most promising and growing field of engineering.

Such advances could one day lead to both biological super-AGI and immortality.

Ciro Santilli is especially excited about DNA-related technologies, because DNA is the centerpiece of biology, and it is programmable.

First, during the 2000's, the cost of DNA sequencing fell to about 1000 USD per genome in the end of the 2010's: Figure 2. "Cost per genome vs Moore's law from 2000 to 2019", largely due to "Illumina's" technology.

The medical consequences of this revolution are still trickling down towards medical applications of 2019, inevitably, but somewhat slowly due to tight privacy control of medical records.

Ciro Santilli predicts that when the 100 dollar mark is reached, every person of the First world will have their genome sequenced, and then medical applications will be closer at hand than ever.

But even 100 dollars is not enough. Sequencing power is like computing power: humankind can never have enough. Sequencing is not a one per person thing. For example, as of 2019 tumors are already being sequenced to help understand and treat them, and scientists/doctors will sequence as many tumor cells as budget allows.

Then, in the 2010's, CRISPR/Cas9 gene editing started opening up the way to actually modifying the genome that we could now see through sequencing.

What's next?

Ciro believes that the next step in the revolution could be could be: de novo DNA synthesis.

This technology could be the key to the one of the ultimate dream of biologists: cheap programmable biology with push-button organism bootstrap!

Just imagine this: at the comfort of your own garage, you take some model organism of interest, maybe start humble with Escherichia coli. Then you modify its DNA to your liking, and upload it to a 3D printer sized machine on your workbench, which automatically synthesizes the DNA, and injects into a bootstrapped cell.

You then make experiments to check if the modified cell achieves your desired new properties, e.g. production of some protein, and if not reiterate, just like a software engineer.

Of course, even if we were able to do the bootstrap, the debugging process then becomes key, as visibility is the key limitation of biology, maybe we need other cheap technologies to come in at that point.

This a place point we see the beauty of evolution the brightest: evolution does not require observability. But it also implies that if your changes to the organism make it less fit, then your mutation will also likely be lost. This has to be one of the considerations done when designing your organism.

Other cool topic include:

It's weird, cells feel a lot like embedded systems: small, complex, hard to observe, and profound.

Ciro is sad that by the time he dies, humanity won't have understood the human brain, maybe not even a measly Escherichia coli... Heck, even key molecular biology events are not yet fully understood, see e.g. transcription regulation.

One of the most exciting aspects of molecular biology technologies is their relatively low entry cost, compared for example to other areas such as fusion energy and quantum computing.

Main section: fusion power.

This is a long haul. But we have to give it a shot.

Ciro Santilli believes that there is a close link between the ability to create disruptive technology, and the desire to find bugs/exploits in systems.

Both of them destabilize society and enterprises.

Some examples:

And yes, this sometimes leads into a fine line between legality and illegality:

Ciro Santilli's own naughty projects are listed at: Section "Ciro Santilli's naughty projects".