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Using CRISPR Technology for Accurate Sample Preparation and Transfection

Using CRISPR Technology for Accurate Sample Preparation and Transfection

Introduction

Clustered Regularly Interspaced Short Palindromic Repeats, or CRISPR for short, became popular in 2012,1 and since then it has become a standard in scientific research. This technique holds great potential for personalized medicine on a level that was not possible before. China is already using the CRISPR technology to conduct clinical trials.

There are many dimensions to the CRISPR technique and these should be carefully considered and executed in order to obtain the required results. But sometimes adoption can precede understanding, as with the case with many other innovative technologies. For example, when the CRISPR technique is used, correct temperatures have to be maintained throughout the duration of sample preparation and transfection but scientists may not be aware of this fact.

What exactly is CRISPR and how does it work?

CRISPR is a gene editing tool which allows highly precise changes to be made inside the DNA. The technique can be applied to all forms of life, from humans to bacteria. It works by using CRISPR associated protein 9 (Cas9) and this detects CRISPR sequences to adhere to a guide RNA whilst at the same time serves as ‘molecular scissors’ to slice the DNA at a preferred site. This process makes it easy to add a new piece of DNA instead of the original sequence.

When compared to other previously existing methods, the specificity of CRISPR/Cas9 provides considerable improvement and as a result, the CRISPR technique has been rapidly and extensively adopted.

Promise and controversy

Researchers across the globe are working hard to discover innovative ways to use the CRISPR technology in the treatment of various diseases, covering from dementia to birth defects. In the US, University of Pennsylvania researchers are set to begin a clinical trial to test the safety and efficacy of CRISPR in humans. However, this field is also fraught with controversy. According to some researchers, CRISPR cannot be used in humans as it is a relatively new technique and its adverse events and potential long-term consequences are not known, while other researchers are concerned about the ethical issues associated with modification of genes in unborn children.

Even the original inventors of CRISPR technology have cautioned against the prospective unethical future applications of the CRISPR technique and recommended a meeting between “experts in law, genetics, and bioethics, and a globally representative team of users and developers of genome engineering technology, in addition to the public, members of the scientific community, and relevant interest groups and government agencies, to consider these critical problems, and where suitable, propose policies.”3

The direct application of the CRISPR technique in human cells has triggered a great deal of interest among the general public. We look forward to the day where Alzheimer’s disease, HIV, and even aging are cured by CRISPR, and at the same time there is a constant worry regarding the slippery slope of the prenatal use of CRISPR, contemplating where the line is between eugenics and medicine. Despite all these hopes and apprehensions, CRISPR also provides a number of indirect potential advantages to human health.

In animal models and cell culture, the use of CRISPR technology has already enhanced preclinical drug discovery and basic research efforts. Toxicity or lack of efficacy contributes to the failure of most clinical trials; for instance, when a drug is given to the mice, they displayed considerable progress without toxicity. This is attributed to the variations between species and represents a major obstacle in the preclinical use of animal models.

With the development of the CRISPR technology, even more accurate cell and animal models of human disease will be made, which can lead to more effective treatments. CRISPR can also be used in plants to create crops that produce greater yield, are healthier, and more resistant to insects and pests without the use of antibiotics.

The value of consistency

With papers exposing considerable challenges in reproducing published data and scientists reporting their own issues in reproducing their peers’ findings, the scientific society has been described as being in a state of “reproducibility crisis.”4,5 This problem also casts doubt on the whole scientific community and erodes the public’s confidence in novel treatments and discoveries. Moreover, it also slows research development, with researchers doubting the validity of research on which their own studies are built upon.

Variations in protocols and reagents between the research groups are the main reason which makes it difficult to replicate that data. Also, within the same laboratory setting, different scientists will not be able to reproduce each other’s data. To achieve considerable improvements, as many variables as possible should be standardized and documented within a procedure.

Innovation solutions

Vitl Life Science Solutions provides breakthrough opportunities to reduce variability in temperatures, both across the experiments and within an experiment. The Ther-Mix and Intelligent Heated Modules are advanced products that provide reproducible and precise temperature settings with user-friendly and programmable interfaces, which enable researchers to generate particular presets for their individual experiments. When performing the CRISPR technique, researchers can use these sophisticated Vitl technologies to avoid major cause of errors in reproducibility and thus enhance transfection efficiency. With the help of Vitl products, researchers can move closer toward the golden age of medicine through CRISPR/Cas9.

The following video shows the application of Ther-Mix and Intelligent Heated Modules:

References

  • Cong, L., Ran, F.A., Cox, D., Lin, S., Barretto, R., Habib, N., Hsu, P.D., Wu, X., Jiang, W., Marraffini, L.A., et al. (2013). Multiplex genome engineering using CRISPR/Cas systems. Science 339, 819–823.

  • S Reardon. First CRISPR clinical trial gets green light from US panel. Nature News. Nature Publishing Group. June 2016. http://www.nature.com/news/first-crispr-clinical-trial-gets-green-light-from-us-panel-1.20137.

  • Baltimore D, Berg P, Botchan M, et al. A prudent path forward for genomic engineering and germline gene modification. Science. 2015;348:36-38. doi:10.1126/science.aab1028.

  • Baker, Monya. “1,500 Scientists Lift the Lid on Reproducibility.” Nature News. Nature Publishing Group. May, 2016.

  • The Open Science Collaboration. “Estimating the Reproducibility of Psychological Science.” Science 349.6251 (2015).

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