Difference between revisions of "Part:BBa K1699002:Design"

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===Design Notes===
 
===Design Notes===
  
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Cloned into pSB1C3 using EcoRI and PstI restriction sites.
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Designed using Benchling to target human UBB gene and to meet registry standards. Synthesized by IDT. Cloned into pSB1C3 using EcoRI and PstI restriction sites.
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<br /> F. tagtcatgGAATTCGCGGCCGCTTCTAG
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<br /> R. tgtatactCTGCAGCGGCCGCTACTAG
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===Source===
 
===Source===
  
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IDT de-novo synthesis.
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Sequence of gRNA: designed using CRISPR tool in Benchling to target human UBB gene.
  
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===References===
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Ribozyme-flanked (RGR) design for the gRNA synthesis under RNA polymerase II promoters:
  
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1. In vivo genome editing using Staphylococcus aureus Cas9.  
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Self-processing of ribozyme-flanked RNAs into guide RNAs in vitro and in vivo for CRISPR-mediated genome editing. Gao Y, Zhao Y. J Integr Plant Biol. 2014 Apr;56(4):343-9. doi: 10.1111/jipb.12152. Epub 2014 Mar 6. http://www.ncbi.nlm.nih.gov/pubmed/24373158
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<br />10.1038/nature14299
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2. Self-processing of ribozyme-flanked RNAs into guide RNAs in vitro and in vivo for CRISPR-mediated genome editing
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Multiplexed and programmable regulation of gene networks with an integrated RNA and CRISPR/Cas toolkit in human cells. Nissim L, Perli SD, Fridkin A, Perez-Pinera P, Lu TK. Mol Cell. 2014 May 22;54(4):698-710. doi: 10.1016/j.molcel.2014.04.022. Epub 2014 May 15. http://www.ncbi.nlm.nih.gov/pubmed/24837679
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<br />http://onlinelibrary.wiley.com/doi/10.1111/jipb.12152/full
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Latest revision as of 12:14, 13 September 2015

gRNA for SaCas9 targeting human ubiquitin B gene


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 146
    Illegal NgoMIV site found at 175
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

Designed using Benchling to target human UBB gene and to meet registry standards. Synthesized by IDT. Cloned into pSB1C3 using EcoRI and PstI restriction sites.

Source

Sequence of gRNA: designed using CRISPR tool in Benchling to target human UBB gene.

Ribozyme-flanked (RGR) design for the gRNA synthesis under RNA polymerase II promoters:

Self-processing of ribozyme-flanked RNAs into guide RNAs in vitro and in vivo for CRISPR-mediated genome editing. Gao Y, Zhao Y. J Integr Plant Biol. 2014 Apr;56(4):343-9. doi: 10.1111/jipb.12152. Epub 2014 Mar 6. http://www.ncbi.nlm.nih.gov/pubmed/24373158

Multiplexed and programmable regulation of gene networks with an integrated RNA and CRISPR/Cas toolkit in human cells. Nissim L, Perli SD, Fridkin A, Perez-Pinera P, Lu TK. Mol Cell. 2014 May 22;54(4):698-710. doi: 10.1016/j.molcel.2014.04.022. Epub 2014 May 15. http://www.ncbi.nlm.nih.gov/pubmed/24837679