Chapter 172 Gene Scissors
After Zhang Wei’s nucleoside modification solved the problem of mRNA breaking through the immune system, the gene editing technology mentioned by Academician Qian became the biggest difficulty in the development of mRNA vaccines.
The difficulty does not mean that it cannot be done, but that it is very difficult to do and requires a lot of money and time.
It is not difficult to find viral antigens, but it was indeed difficult to edit the gene sequence corresponding to this antigen separately based on the genetic technology at the time.
For example, the target antigen of the new coronavirus is the spike protein of the virus, so as long as the mRNA sequence of the spike protein is cut out and used as a template, the required mRNA stock solution can be obtained to prepare the corresponding of mRNA vaccines.
However, how to cut the mRNA sequence of this spike protein from the virus gene requires a lot of complicated experiments, and whether it can succeed in the end depends on some luck.
Humanity discovered restriction endonuclease in the 1970s. This enzyme can recognize specific deoxyribonucleotide sequences and target the gap between two deoxyribonucleotides at a specific site. Phosphodiester bonds are cleaved.
In human terms, this enzyme can cut gene fragments.
Since then, humans have had gene editing technology.
However, using endonuclease to cut gene fragments has a fatal disadvantage, that is, the accuracy and efficiency are very low.
The first is that endonucleases are often unable to enter target cells, which means that when I want to cut gene fragments, I cut more than a dozen times, but none of them hit the mark.
The second reason is that the accuracy of endonuclease is too poor. For example, what I want to cut is this gene fragment, but in the end what is cut is the other end of the gene fragment, or it is too much or too little. .
Whether it is impossible to cut or the cut is inaccurate, it is something that greatly affects efficiency. What's more, the cost of using this method to edit genes is quite high. Unless you have a mine at home, you can't afford it!
Gene editing technology in the past few decades has also relied on restriction endonucleases for research. All gene editing research has tried to improve the accuracy of endonucleases.
In the mid-1980s, scientists discovered zinc finger proteins, which can be linked to endonucleases, and then developed zinc finger nuclease technology. This technology can achieve targeted cutting of DNA and greatly improve the accuracy.
Until the first decade of the 21st century, humans used this gene editing technology.
But this technology also has great limitations. First of all, zinc finger proteins can affect the recognition and binding of specific nucleotide sequences, which means that you can't cut whatever you want.
The second thing is that in human genes, every 500 bases can have a target for cutting. In other words, you can’t cut as long as you want. You need 500 bases to make one cut. .
If you want a 600-base fragment, you can only cut the 1,000-base fragment. The 600-base fragment cannot be cut with one knife.
Finally, there is the price factor. Zinc finger proteins are very expensive. One experiment costs almost US$5,000 for zinc finger proteins. Even Americans with deep pockets cannot afford such experiments.
If this kind of gene editing technology is used in the development of mRNA vaccines, hundreds of experiments will be carried out and the gene sequence corresponding to the antigen will not be obtained. It is equivalent to spending millions of dollars and not even getting any results. See.
Compared with the development process of inactivated vaccines, the cost is definitely advantageous. The technology of virus inactivation has been used for decades, it is very mature, and the cost is completely controllable. within.
Considering this aspect, Sinopharm’s unwillingness to spend money on mRNA vaccine technology is also a well-founded decision.
……
Academician Qian explained very clearly, and the people present were all professionals who could understand the concerns of Sinopharm Group.
Zhang Wei did not give up, he said: "The currently used gene editing mainly uses zinc finger nuclease technology. Although this technology has been used for twenty years, it does have many shortcomings, which will increase the cost of mRNA vaccine development. .
If I can improve the gene editing technology, or develop a new gene editing technology to reduce the cost of research and development of mRNA vaccines, can Sinopharm consider investing in the research and development of mRNA vaccines? ”
“You want to develop new gene editing technology?” The three academicians were surprised at the same time.
Meng Heming also quickly advised: "Zhang Wei, gene editing technology cannot be developed by research and development. The existing zinc finger nuclease technology has been used for more than 20 years, and there is no new one. With the emergence of technology, this kind of matter needs to be considered in the long term, and you can't make big claims!"
"Don't worry, Director Meng, since I dare to mention this matter, I must have an idea," Zhang Wei said calmly. .
The three academicians also felt that Zhang Wei was bragging, but seeing Zhang Wei's calm look, they felt that Zhang Wei might not be innocent.
Considering that Zhang Wei is a genius scholar who was able to publish an article in "Science" at the age of twenty, maybe he really has a way to develop new gene editing technology.
The three academicians looked at each other, and finally it was Academician Qian who asked: "Teacher Zhang Wei, the new idea of gene editing technology you mentioned, is it the use of transcription activators to activate The transcription ability of genes, right?”
“Academician Qian has also read the literature in this area?” Zhang Wei asked with a smile.
“I have read some literature, and there has been research in this area abroad.” Academician Qian nodded, and then explained to Director Wang; “There is a kind of Xanthomonas, which is a common pathogen in plants. Bacteria are also the main cause of rice bacterial blight
The Xanthomonas bacterium secretes a protein, a transcription factor, that binds to the host genome and activates its ability to transcribe.
So some scientists hope to use this transcription ability to activate gene expression to achieve the purpose of gene editing. If this transcription factor can be fused with an endonuclease, it will bring about a revolutionary gene editing technology. "
Director Wang nodded, roughly understanding it, and then he asked: "This project shouldn't be easy to implement, right? "
"Of course it is not simple. I have only mentioned it in some foreign literature. There may be several teams conducting research in this area with the funding of large foreign companies. There should be no one in China. Do research in this area. "Academician Qian said.
The next second, everyone looked at Zhang Wei, as if asking, can you really succeed in such a difficult project?
Zhang Wei was still calm, and he said: "Since I brought it up, I must be sure."
"It's true that a newborn is not afraid of tigers. I was like Teacher Zhang Wei when I was so young. , I am full of enthusiasm! But scientific research cannot be done just by talking about it!" Academician Qian reminded with a smile.
"How about this? Give me half a year. After half a year, I will ask Director Wang and three academicians to come again to see my results. If the results can really catch your eyes by then , then Sinopharm should no longer refuse the mRNA vaccine project," Zhang Wei said.
"Half a year? Is it enough time?" The person who asked the question this time was Director Wang. He continued: "Although I am not a scientist, I also know that scientific research cannot be rushed, and experiments also need to be done in a hurry. Time.
Teacher Zhang Wei, it’s better to set the time more loosely! Anyway, it will take more than ten years to develop a vaccine, so there is no rush.”
"Director Wang, don't worry. The biggest characteristic of me doing scientific research is that I am fast! I said results will be produced in half a year, and results will be produced in half a year! If you don't believe it, I can issue a military order!" Zhang Wei said confidently. "It's not necessary. Our place is not a military camp. Why should we issue a military order?" Director Wang quickly waved his hand.
For this kind of thing, even if it is a military order, there is no basis for punishment. Sinopharm and Tsingta University are two different units, and they cannot even punish them with three drinks.
Even if Director Wang thought Zhang Wei was bragging, he could only be humane, put on an expectant expression, and said, "Then I'm looking forward to Mr. Zhang Wei's research and development results."< br>
……
After seeing off the three academicians, Zhang Wei was called to the office by Meng Heming.
"Are you sure you want to do this project? I have just checked some information. The idea of using transcription activators for gene editing is certainly feasible, but it is currently only at a theoretical level." Meng Heming said.
“I did it, so it’s not a theoretical level.” Zhang Wei said confidently.
“This project is very difficult!” Meng Heming then reminded.
“How can we trap Sinopharm without releasing a big satellite? We have to let them invest in the research and development of mRNA vaccines!”
"Actually, you don't have to worry about the research and development of mRNA vaccines. Your paper has just been published. Those pharmaceutical companies are still in the wait-and-see stage. It is normal that no companies will invest."
"Can't you be in a hurry? I I still want to win the Nobel Prize soon! Who is the youngest Nobel Prize winner? Lawrence Bragg! He won the Nobel Prize at the age of 25. I am already twenty, and I still have five years to go! He said with a smile.
"Lawrence Bragg won the Nobel Prize in 1915! It was still the barbaric age of science, how can it be compared with now!"
Meng Heming paused for a moment, then continued: "Again Come on, you don’t have to spend money to do scientific research? Where do you have to apply for it?
“Don’t you still have a balance for the nucleoside modification project? You were all staring at my balance yesterday, and now you have forgotten about it? I also happened to use this new project to spend all the balance. Well, you won’t have to worry about it then!” Zhang Wei responded.
"What you want to do now is gene editing. How many experiments can you do with your little balance? Not to mention other things, such as the existing zinc finger nuclease technology, the cost of one experiment is about US$5,000. ”
“The transcription activator is secreted by Xanthomonas. Xanthomonas is very common. You can probably collect it from any withered grass leaves on the roadside. The cost is not that high!” Zhang Wei! He still had a cheerful expression.
“But endonuclease must be used, right? Other reagents must be used, too? Maybe halfway through your experiment, you will find that you need some expensive reagent to proceed!”
"It's okay, I'll apply to the school then! I went to blackmail the principal, but he was embarrassed not to give it? Our Tsinghua University has billions of scientific research funds a year, why should I spend more!" Zhang Wei Said heartlessly.
“How can you spend billions of dollars a year on you? No one else is doing projects?”
"Can projects done by other people win Nobel Prizes?"
Zhang Wei asked back, and Meng Heming was immediately speechless.
You can win the Nobel Prize, you are awesome!
Zhang Wei still had a smile on his face and said: "Director Meng, let me do it first and give it a try. You know that I am lazy and cowardly. If it comes later, If the project cannot continue, you don’t need to ask, I will stop it myself!
But if I develop a new gene editing technology, it will be a major scientific research achievement for the entire biological sciences. The research city will set off a revolution, and our Tsinghua University will make a lot of money just by selling patents!”
...
The zinc finger nuclease technology, developed in the 1980s and used ever since, was the first generation of gene editing technology and was also a commonly used technology at the time.
The transcription activator mentioned by Academician Qian is the second generation of gene editing technology. This technology was only developed around 2010.
This technology is also developed based on endonuclease. Due to the cumbersome construction process, the operation is much more difficult than the first generation technology. It requires operators with certain experimental skills to implement it. operate.
Furthermore, this second-generation gene editing technology is relatively easy to miss the target. In actual operation, the efficiency does not exceed 25%. In other words, when performing gene editing, only one hit can be achieved after every four cuts.
Such efficiency is actually very worrying when it comes to conducting experiments. This means that the experimenter has to prepare four samples to ensure that one sample can hit the target cell.
If you are less lucky, each portion will have a 75% chance of missing the target, which is not impossible! After all, the probability of all four samples missing the target is as high as 31.64%. Eight samples must be prepared to ensure a 90% probability of hitting the target cells.
Compared with the first generation of gene editing technology, this second generation technology is already a revolutionary leap, but this is not Zhang Wei’s goal.
When Academician Qian mentioned transcription activators, Zhang Wei also dropped the smoke bomb.
What Zhang Wei wants to do is actually the third generation of gene editing technology.
That is CRISPR-Cas, commonly known as “gene scissors”!
The first two generations of gene editing technology were all based on endonucleases, but the third generation of genetic scissors is different. It is a brand-new gene editing theory.
In the genome of bacteria, there is a repetitive sequence called CRISPR. This sequence is very ancient and can be traced back to the birth of bacteria.
This sequence is the immune weapon used to fight bacteria and viruses.
When bacteria detect a viral invasion, CRISPR will generate an enzyme called Cas9, forming an immune system called CRISPR-Cas9.
This immune system can directly excise the DNA sequence of the invading virus, thereby resisting viral invasion.
What we normally know about the immune system is that it produces antibodies based on antigens and then exerts an immune effect.
This kind of CRISPR-Cas9 directly targets the DNA of the invader, directly chops the DNA of the invader, and naturally eliminates the invader in the bud.
This is exactly the principle of genetic scissors. Since CRISPR-Cas9 directly cuts the DNA sequence, it means that this technology can be used to cut gene fragments.
I want that gene, just use CRISPR-Cas9 to cut it, and everything will be fine!
(End of this chapter)