Device identifies genetic mutations using a CRISPR-chip made from graphene

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CRISPR-chip Graphene
CRISPR-Chip uses deactivated CRISPR-Cas9 protein, tethered to a transistor made of graphene, to detect specific genetic sequences in a DNA sample.

A portable device to diagnose hereditary anomalies

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team of engineers hopes a brand-new device called the CRISPR-Chip might swiftly diagnose hereditary anomalies without requiring to send information to a lab.

Genetics has gotten in unchartered territory at a scorching rate over the past couple of years, and currently, a group of scientists from UC Berkeley wants to take things a step further with a brand-new tool that could soon remain in various physicians’ offices.

Imagine a tiny robot that was programmed to unzip the DNA double helix, search through the whole genome, identify a match to any sequence you’d like, and if you asked it to, it could then further edit the genome to your liking

Called the CRISPR-Chip, the tool integrates gene-editing methods of CRISPR with electronic transistors made from the ‘Marvel product’ graphene. According to its designers, the portable device could be used to detect genetic diseases or to review the precision of gene-editing techniques in just a few minutes.

“We have created the initial transistor that utilizes CRISPR to search your genome for possible mutations,” stated Kiana Aran, who initially developed the concept for the device. “You just place your cleansed DNA sample on the chip, enable CRISPR to do the search and the graphene transistor reports the outcome of this search in minutes.” 

Graphene Crispr-chip
CRISPR-Chip uses deactivated CRISPR-Cas9 protein, tethered to a transistor made of graphene, to detect specific genetic sequences in a DNA sample. (Graphic courtesy Kiana Aran)

The CRISPR-Chip uses graphene to report CRISP activity

Publishing the results in Nature Biomedical Engineering, the team stated that unlike the majority of forms of genetic testing, the CRISPR-Chip makes use of nanoelectronics to spot any type of mutations in DNA samples without ‘intensifying’ or duplicating the DNA section millions of times over in a procedure referred to as polymerase chain reaction (PCR).

This indicates that the specimen does not need to be gone back to a laboratory – which incurs great expenses, both money and time – but can be analyzed in a doctor’s office.

Describing the science behind it, the team said that usually, the Cas9 protein that permits CRISPR to operate needs a large fragment of ‘guide RNA’. The protein first unzips the double-stranded DNA and also scans through until it finds the sequence that matches the guide RNA it intends to cut, and afterward latches on.

Our vision is to use technological advancement in the digital world and build tools to provide us with instant access to the world of biology

Optical methods don’t have the speed to follow the Cas9 activity quickly or need the incorporation of optical tags and/or amplification. Silicon-based electronic devices do not have the level of sensitivity needed for spotting the task of CRISPR activity, as well as silicon does not play great with biological material either. It was required a sensitive electronically active biocompatible product: Graphene.

Graphene is one of the most popular nanomaterials. It is a sheet of one atom thick carbon and also the very first two-dimensional semiconductor discovered. It has features like high electron mobility and also chemical stability in salt water that make it an excellent electrical channel for reporting the activity of our biological sample.

CRISPR dCas9 variant hangs on to a specific DNA sequence

To develop this system, the UC Berkeley team utilized a changed variation of Cas9 protein (dCas9) within the CRISPR complex to ensure that when it discovers the matching DNA sequence to the guide RNA, it holds on instead of doing a cut to edit. Thousands of these neutered Cas9 enzymes are placed on each graphene sensing unit using special chemical linkers. The CRISPR system remained active, which took a while as well as the job of most of the lab participants to optimize. With an active dCas9 on the graphene surface area, it allows to pick up the change in energy that happens at the surface of the graphene when the CRISPR hangs on to the part of the genome it was designed to find, informing the CRISPR-Chip device when it finds a ‘hit’

Kiana Aran CRISPR-chip
Aran conceived of CRISPR-Chip while a postdoctoral scholar at UC Berkeley and developed it as an assistant professor at KGI.

The ultimate objective of the group is to ‘multiplex’ the device, enabling doctors to connect to several guide RNAs at once to simultaneously find several genetic anomalies in just a few minutes.

“Envision a web page with a lot of search boxes – in our case, transistors – and you have your overview RNA info in these search boxes, as well as each of these transistors will do the search and report the outcome digitally,” Aran said.

The CRISPR-Chip can be used to identify efficiency and safety of CRISPR gene editing

For the research study to show its merit, the group used it to discover two usual genetic anomalies in blood samples from Duchenne muscular dystrophy (DMD) patients. If confirmed effective, the CRISPR-Chip would be particularly useful for DMD screening as it is presently expensive and time-consuming to check for the extreme muscle-wasting illness.

Irina Conboy, a co-author of the study, clarified: “With an electronic device, you could make guide RNAs throughout the entire dystrophin genetics, and afterward you might simply evaluate the entire series of the gene in a matter of hours”.

“You could evaluate moms and dads, and even newborns, for the visibility or lack of dystrophin mutations – and after that, if the anomaly is discovered, therapy could be started early, before the condition has formed.”.

We merely need to have tools that can supply a much deeper level of insight before we are successful in ourselves with human genome editing. Therefore, we are eager to develop CRISPR-Chip for CRISPR quality control (CRISPR QC) for evaluating “on-target vs. off-target” and for gRNA validation

However, diagnostics is not the only application of the CRISPR-Chip. As an example, the CRISPR-Chip can be used to enhance the efficiency and safety of CRISPR gene editing. Validation in vitro research studies is often needed to optimize CRISPR effectiveness. However, these studies primarily assess CRISPR effectiveness at the prompt location of the target.

Off-target CRISPR activities, as it can edit unexpected locations in the genome, is a real issue we need to reduce at any expense as we approach CRISPR for therapeutic usage. Undesirable CRISPR editing, including deletions as well as alterations that occur much from the targeted genes, could cause major clinical outcomes. Furthermore, most efficient gRNAs, that are usually picked after gRNA recognition research studies, may not do the same in-vivo due to the complexity of the genome which differs from the naked DNA usually used in validation studies.

Source

Reza Hajian, Sarah Balderston, Thanhtra Tran, Tara deBoer, Jessy Etienne, Mandeep Sandhu, Noreen A. Wauford, Jing-Yi Chung, Jolie Nokes, Mitre Athaiya, Jacobo Paredes, Regis Peytavi, Brett Goldsmith, Niren Murthy, Irina M. Conboy, Kiana Aran. Detection of unamplified target genes via CRISPR–Cas9 immobilized on a graphene field-effect transistor. Nature Biomedical Engineering. Published online Mar 25, 2019. doi: 10.1038/s41551-019-0371-x.