CRISPR Cas9 is a powerful gene editing tool that cuts the DNA in a precise manner. The most common use of CRISPR Screening is finding gene targets for various diseases. The process of a typical CRISPR Screening experiment is as follows :
Why not MAGeCK?
There are various tools developed for processing CRISPR Screening data. Some of them are MAGeCK, HitSelect, ScreenBEAM, BAGEL, sgRSEA, PBNPA, MAGeCK VISPR which use different algorithms. All these tools are script based which can be used only by bioinformaticians or tech-savvy biologists. These are not friendly to most scientists. Apart from this, there were some other challenges as well that were faced by scientists and even bioinformaticians. Let’s try to understand the challenges taking the example of one of the most widely used tools for CRISPR Screening- MAGeCK.
- Command line interface: It’s a command-based tool that requires a specific set of commands to be typed in order to process CRISPR screening data. Not everyone is comfortable with a terminal/command prompt
- Limited processing power: CRISPR screening datasets are huge, in GBs. Processing that data with the computational power of just a local computer takes hours. Due to the limitation of computational power, only one comparison can be processed at a time. The computer needs to stay on throughout the time, analysis is running and the output gets generated only there
- No dashboard: Output generated is in the form of text files which are not very convenient for interpretation
- No quality check: There’s no quality check module which can give a sense of the quality of input CRISPR screening data
Why not MAGeCK VISPR?
MAGeCK VISPR was developed in order to overcome some of the shortcomings of MAGeCK. It uses the base algorithm of MAGeCK and does everything that MAGeCK does. On top of that, it has some additional algorithms and features that make it much more useful than MAGeCK. Those features are:
- Quality check: This module lets us identify the outliers in the CRISPR screening dataset which we can remove and rerun the analysis
- Visualization dashboard: The output is in the form of an interactive visual dashboard which makes it much easier to interpret
Even though MAGeCK VISPR was a significant improvement over MAGeCK, there were some critical limitations that persisted.
- No multiple comparison dashboard: There was still no way to look at multiple comparisons on the same dashboard. This feature has tremendous value to a CRISPR screening study. Scientists often find a target and validate their target over multiple experiments. MAGeCK VISPR did not give an easy way to do that
- Command line interface: It was still a command based tool
- Limited processing power: Used computational power of a local computer
- No customizability: The analysis could not be customized
- No restore analysis: Previous analysis could not be recalled. The analysis needed to be run again to have a look at the dashboard
Why Polly CRISPR Screening?
Polly CRISPR Screening, based on MAGeCK algorithm, is a cloud-based tool with an interactive interface instead of a command line interface. This decreases the learning curve of the tool and makes it much easier to use. And, considering it is cloud-based, there’s no need to install anything on the local computer and data can be processed from any computer.
The benefits of using Polly CRISPR screening workflow are:
- Fast processing: Processing is up to 2x faster compared to a local computer. (Specifications for comparison: Intel Core i5 with 2.5GHz frequency, 4 GB RAM)
- Multiple comparisons: Polly CRISPR screening workflow lets you run multiple comparisons at the same time, processing all of them simultaneously
- Restore analysis: The output is stored on the cloud which can be restored later by logging into your Polly account
- Share and collaborate: You can share your project with peers and collaborate in studies with them over the platform
- Multiple comparisons dashboard: This dashboard provides information about how genes behave across different comparisons. This helps to identify whether a specific target is consistent across various analyses and experiments helping validate or reject a hypothesis much faster. The dashboard also has an option to generate a list of all the genes that are common across multiple experiments. It helps in identifying gene targets
- sgRNA removal: There are sgRNAs which have an abruptly high count in a sample. The sgRNA remove function helps to remove such gRNAs from the experiment and re-run the MAGeCK analysis on the dashboard
- IPython notebook: You can customize the workflow by writing custom scripts in the IPython notebook feature. This is particularly useful as every lab has its own workflow for experiments they conduct
We hope to add more features to the tool to make it even more useful. One such key feature we believe would add a lot of value is giving related publications to the ongoing study. Suppose you find that gene “XYZ” is of some importance to disease “ABC”. You can get a list of publications which relate the mentioned disease to the mentioned gene. This makes it much faster to accept or reject the hypothesis.
You read how Polly CRISPR screening workflow has a huge edge over anything else available for CRISPR screening studies. Now it is time you check it out yourself.
To experience Polly CRISPR Screening Workflow, book a quick demo here.
Also published on Medium