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CRISPR-CBEI

a web-based designing tool forCytosineBaseEditor mediated geneInactivation

1.Input
Three steps for CBEI prediction:
1.Enter the sequence in Fasta format and set parameters about intron and ORF identification.Click'Submit'to identify ORFs.Although the software supports multiple Fasta sequences,we really recommend that enter only one.Try[Demo]
2.Select the ORF that is expected to be predicted by CBEI and click'Predict'.
3.View or export CBEI-predict results.
Sequence
ORF settings
Note:
Introns can be customized and the CrisprCBEIdoes not consider alternative splicing currently.If set up,the space between introns and exons will also be considered in subsequent CBEI predictions.

The ORF Finder we provided is for basic ORF recognition and is not suitable for new gene prediction.The software searches the six reading frames(+1,+2,+3,-1,-2,-3)for ORF with ATG or others as initiation codons,TAG,TAA,and TGA as termination codons.
Intron
Genetic code
Start codon
Stop codon
Minimum length
ORF identification
2.ORF detection
All detected ORF sequences are displayed. Click ORF in the diagram or table for the next CBEI design.
3.CBEI design
ORF info
Diagram
Base editor
You can customize the base editor by changing the following parameters.If you need to add custom content,please contact us.
PAM
Spacer length
Spacer location
Editing window
Search region
CBEI-PredictRe-select ORFReset CBEI settingsReset all
4.Result summary
  • 1.Potential editing sites[Details]
    • 2.Statistics
      • 2.1 Editable site statistic
      • 2.2 Distribution of editable site
    Result detailsReset CBEI settingsReset all
    5.Result details
    • 1.Click on the"+"in the first column of each row for a more detailed prediction.
    • 2.Select the interested spacer sequences and click"off-target prediction"to make off-target prediction.
    1.Settings
    Three steps for off-target prediction:
    1.Enter the spacer sequences of plain text format(one spacer per line),and the recommended length is 15nt-25nt.Try[Demo]
    2.Select the genome file in Fasta format(genome file size is not limited,but the large file may reduce the rate of the off-target prediction,please see the page of‘Help’for details).
    3.Set other parameters and click‘Predict!’.
    Spacers
    Diagram
    Base editors
    You can customize the base editor by changing the following parameters.If you need to add custom content,please contact us.
    Note,'Spacer length'and'Edit window'bellow are only used for drawing and do not participate in off-target calculation.
    PAM
    Spacer length
    Spacer location
    Editing window
    Genome.fa [Demo]
    Mismatch<=
    2.Off-target prediction result
    Crispr-CBEI update log
    Update time Log
    2018-10-28 Release Crispr-CBEI.
    2018-12-27 Add localy off-target prediction function.
    2019-08-19 Rearrange UI,prediction logic and add ORF identification.
    2019-09-01 The Python version of CrisprCBEI released (autocbei).
    2019-10-27 Increased support for introns.
    2020-06-30 "autocbei" supported "pip" and "conda" installations.
    Shiheng Tao Lab
    Address
    College of Life Sciences and State Key Laboratory of Crop Stress Biology in Arid Areas
    Bioinformatics Center
    Northwest A&F University
    Yangling,Shaanxi,China,712100
    Contact
    Tel:(+86-029)87091060
    Mail:shihengt@nwsuaf.edu.cn
    Quanjiang Ji La
    Address
    School of Physical Science and Technology
    ShanghaiTech university
    Shanghai,China,201210
    Contact
    Mail:quanjiangji@shanghaitech.edu.cn
    Website BUGs report
    Haopeng Yu
    Mail:atlasbioin4@gmail.com
    Zhaowei Wu
    Mail:wuzw1@shanghaitech.edu.cn
    1.User manual

    We have prepared a detailed, step-by-step user manual for CBEI design or off-target prediction.

    Please download and view.

    2.How to set introns

    When ORF detection is performed, you can set whether the input sequence contains introns in "Customize ORF detection". At this point, you can enter the coordinates of the CDS. Separate exons with "," and use ".." to separate the begin and end of exons, as in: 1..102, 216..467

    You can search for related genes in NCBI, GeneBank format will mark the location of CDS, and copy it directly. How to set up exons is clearly explained in the user manual.

    3.Off-target prediction

    CRISPR-CBEI supports flexible local off-target prediction,that is,you can select any desired local Fasta file to compare to the spacer sequence chosen! If you used our supplied Demo sequence,the LacZ gene,you could download this test genome sequence, Test.fa ,for off-target prediction.

    Select your Fasta file (e.g.,Test.fa),and then select "Mismatch" number(Default: mismatch is less than or equal to 3),click "Start prediction!" ,CRISPR-CBEI will quickly complete the calculation subsequently and display the predicted result. Of course,you can use cDNA,ORF or CDS sequences in Fasta format to make the off-target prediction.

    4. The time consumed in the off-target prediction.

    CRISPR-CBEI off-target prediction function is a front-end prediction tool that supports local Fasta format files for off-target prediction without uploading to the server. Through algorithm optimization, CRISPR-CBEI does not limit the size of Fasta files, and the spacer alignment speed is fast. The whole off-target calculation process is asynchronous and does not occupy much memory (about 200MB), so it would not affect other applications of the computer. Computational efficiency depends on the CPU, and the higher the primary frequency, the higher the efficiency. Note that, in our tests, we also found that running other computing tasks on the computer at the same time increased the time consumption.

    Test environment: i5-3470 3.20GHz

    Species File size Mismatch<=3 Mismatch<=2 Mismatch<=1 Mismatch==0
    5. Abbreviations used
    • CBEI : Cytosine Base Editor mediated gene Inactivation.
    • PAM : Protospacer-adjacent motif. If you have additional PAM mode requirements, please contact us.
    • Strand : The positive (+) or the negative (-) strand in which the PAM is located.
    • Edit window : The potential edit region for CRISPR/Cas9-guided cytosine deaminase was included in the bracket, their adjacent codons were also showed and separated by commas.
    • Edit codon : The potential codon of the editable cytosine.
    • Edit nucleotide position : The position of the potential edit base.
    • Edit pattern : The edit pattern indicated the adjacent nucleotide at 5’ of the editable cytosine. Typically, the in vitro activity of the base editors follows TC ≥ CC ≥ AC > GC.
    • Edit location in gene : The relative position of the potential edit base in CDS.