“CRISPR” stands for Clustered Regularly Interspaced Short Palindromic Repeats, which are the hallmark of a bacterial defense system which forms the basis for the popular CRISPR-Cas9 genome editing technology.
CRISPR-Cas9 is a genome editing tool that is creating a buzz in the science world. It is faster, cheaper and more accurate than previous techniques of editing DNA and has a wide range of potential applications.
Through a few clever molecular hacks, researchers at Columbia UniversityMedical Center have converted a natural bacterial immune system into a microscopic data recorder, laying the groundwork for a new class of technologies that use bacterial cells for everything from disease diagnosis to environmental monitoring.
The researchers modified an ordinary laboratory strain of the ubiquitous human gut microbe Escherichia coli, enabling the bacteria to not only record their interactions with the environment but also time-stamp the events.
“Such bacteria, swallowed by a patient, might be able to record the changes they experience through the whole digestive tract, yielding an unprecedented view of previously inaccessible phenomena,” says Harris Wang, assistant professor in the Departments of Pathology & Cell Biology and Systems Biology at CUMC and senior author on the new work, described in today’s issue of Science.
Other applications could include environmental sensing and basic studies in ecology and microbiology, where bacteria could monitor otherwise invisible changes without disrupting their surroundings.
Wang and members of his laboratory created the microscopic data recorder by taking advantage of CRISPR-Cas, an immune system in many species of bacteria. CRISPR-Cas copies snippets of DNA from invading viruses so that subsequent generations of bacteria can repel these pathogens more effectively.
As a result, the CRISPR locus of the bacterial genome accumulates a chronological record of the bacterial viruses that it and its ancestors have survived. When those same viruses try to infect again, the CRISPR-Cas system can recognize and eliminate them.
“The CRISPR-Cas system is a natural biological memory device,” says Wang. “From an engineering perspective that’s actually quite nice, because it’s already a system that has been honed through evolution to be really great at storing information.”
CRISPR-Cas normally uses its recorded sequences to detect and cut the DNA of incoming phages. The specificity of this DNA cutting activity has made CRISPR-Cas the darling of gene therapy researchers, who have modified it to make precise changes in the genomes of cultured cells, laboratory animals, and even humans. Indeed, over a dozen clinical trials are now underway to treat various diseases through CRISPR-Cas gene therapy.
But Ravi Sheth, a graduate student in Wang’s laboratory, saw unrealized potential in CRISPR-Cas’s recording function. “When you think about recording temporally changing signals with electronics, or an audio recording … that’s a very powerful technology, but we were thinking how can you scale this to living cells themselves?” says Sheth.
To build their microscopic recorder, Sheth and other members of the Wang lab modified a piece of DNA called a plasmid, giving it the ability to create more copies of itself in the bacterial cell in response to an external signal. A separate recording plasmid, which drives the recorder and marks time, expresses components of the CRISPR-Cas system. In the absence of an external signal, only the recording plasmid is active, and the cell adds copies of a spacer sequence to the CRISPR locus in its genome.
When an external signal is detected by the cell, the other plasmid is also activated, leading to insertion of its sequences instead. The result is a mixture of background sequences that record time and signal sequences that change depending on the cell’s environment. The researchers can then examine the bacterial CRISPR locus and use computational tools to read the recording and its timing.
The current paper proves the system can handle at least three simultaneous signals and record for days.
“Now we’re planning to look at various markers that might be altered under changes in natural or disease states, in the gastrointestinal system or elsewhere,” says Dr. Wang.
Synthetic biologists have previously used CRISPR to store poems, books, and images in DNA, but this is the first time CRISPR has been used to record cellular activity and the timing of those events.
Chinese scientists have created low-fat pigs using new genetic engineering techniques. In a paper published in the Proceedings of the National Academy of Sciences, the scientists report that they have created 12 healthy pigs with about 24 percent less body fat than normal pigs.
The scientists created low-fat pigs in the hopes of providing pig farmers with animals that would be less expensive to raise and would suffer less in cold weather.
The animals have less body fat because they have a gene that allows them to regulate their body temperatures better by burning fat. That could save farmers millions of dollars in heating and feeding costs, as well as prevent millions of piglets from suffering and dying in cold weather.
The Chinese scientists created the animals using a new gene-editing technique known as CRISPR-Cas9. It enables scientists to make changes in DNA much more easily and precisely than ever before.
Pigs lack a gene, called UPC1, which most other mammals have. The gene helps animals regulate their body temperatures in cold temperatures. The scientists edited a mouse version of the gene into pig cells. They then used those cells to create many cloned pig embryos.
Next, scientists implanted the genetically modified cloned pig embryos into 13 female pigs. Three of the female surrogate mother pigs became pregnant, producing 12 male piglets.
For the first time, a primitive movie has been encoded in – and then played back from – DNA in living cells.
The research team shows in foundational proof-of-principle experiments that the CRISPR system is able to encode information as complex as a digitized image of a human hand, reminiscent of some of the first paintings drawn on cave walls by early humans, and a sequence of one of the first motion pictures made ever, that of a galloping horse, in living cells.
When a virus infects a bacterium, CRISPR cuts out part of the foreign DNA and stores it in the bacteria’s own genome. The bacterium then uses the stored DNA to recognize the virus and defend against future attacks.
The researchers translated five frames from the race horse in motion photo sequence into DNA. Over the course of five days, they sequentially treated bacteria with a frame of translated DNA. Afterwards, they were able to reconstruct the movie with 90 percent accuracy by sequencing the bacterial DNA.
Although this technology could be used in a variety of ways, the researchers ultimately hope to use it to study the brain.
Scientists say it is a major step toward a “molecular recorder” that may someday make it possible to get read-outs, for example, of the changing internal states of neurons as they develop.
The CRISPR system helps bacteria to develop immunity against the constant onslaught of viruses in their different environments. As a memory of survived infections, it captures viral DNA molecules and generates short so-called “spacer” sequences from them, that are added as new elements upstream of previous elements in a growing array located in the CRISPR locus of bacterial genomes.
The by now famous CRISPR-Cas9 protein constantly resorts to this memory to destroy the same viruses when they return. Besides Cas9, which has become a widely used genome engineering tool, other parts of the CRISPR system, however, have so far not been exploited much technologically.