CRISPR Featuring IDT Products and Services

gBlocks® Gene Fragments are synthetic high-fidelity, double-stranded DNA that are ideal for creating sgRNA constructs for targeting the Cas9 endonuclease in CRISPR applications. gBlocks Gene Fragments are also ideal for assembling new, codon optimized Cas9 variants. The following list of citations provides specific examples of the use of gBlocks Gene Fragments in CRISPR research.

  1. Cas9 ribonucleoprotein delivery via microfluidic cell-deformation chip for human T-Cell genome editing and immunotherapy
    Han X, Liu Z, et al.
    Advanced Biosystems, 1(1):[Epub ahead of print] (2017)

    Here, researchers deliver Cas9 ribonucleoprotein (RNP) to various cell types, including human primary CD4+ T cells, via a novel microfluidic cell deformation-based method.

  2. Gene editing in mouse zygotes using the CRISPR/Cas9 system
    Wefers B, Bashir S, et al.
    Methods, S1046-2023(16):[Epub ahead of print] (2017)
    This publication details the process of designing knock-out and knock-in CRISPR experiments for the generation of new mouse mutants. It outlines proper preparation of Cas9 ribonucleoprotein, as well as procedures for delivering the complex to mouse zygotes by way of microinjection and electroporation.
  3. Insertional mutagenesis by CRISPR/Cas9 ribonucleoprotein gene editing in cells targeted for point mutation repair directed by short single-stranded DNA oligonucleotides
    Rivera-Torres N, Banas K, et al.
    PLoS One, 12(1):e0169350 (2017)
    This publication from the laboratory of Dr. Eric Kmiec highlights the advantages of using CRISPR-Cas9 ribonucleoprotein for DNA cleavage along with single-stranded DNA oligonucleotides for repair of single base mutations, and examines the mechanism of repair in greater detail.
  4. Markerless chromosomal gene deletion in Clostridium beijerinckii using CRISPR/Cas9 system
    Wang Y, Zhang ZT, et al
    J Biotechnol, [Epub ahead of print]: (2015)
  5. High-efficiency multiplex genome editing of Streptomyces species using an engineered CRISPR/Cas system
    Cobb RE1, Wang Y, Zhao H.
    ACS Synth Biol, [Epub ahead of print]: (2014)
  6. The Mitochondrial Translocator Protein (TSPO) Inhibits HIV-1 Envelope Glycoprotein Biosynthesis via the Endoplasmic Reticulum (ER)-Associated Protein Degradation (ERAD) Pathway.
    Zhou T, Dang Y, Zheng YH.
    J Virol, 88(6):3474–3484 (2014)
    A gBlocks Gene Fragment was used to generate a single-guide RNA for use with the CRISPR/Cas9 system. The 455 bp fragment contained a U6 promoter, target sequence, and guide RNA scaffold.
  7. CRISPR/Cas9-mediated genome modification in the mollusc, Crepidula fornicata
    Perry KJ, Henry JQ
    Genesis, 53(2):237–244 (2014)
    A gBlocks Gene Fragment is used to create a donor DNA sequence for homologous DNA repair in a CRISPR/Cas9 genome modification protocol
  8. Fanconi Anemia Gene Editing by the CRISPR/Cas9 System
    Osborn MJ, Gabriel R, et al
    Hum Gene Ther, 26(2):114–126 (2014)
  9. Highly efficient targeted chromosome deletions using CRISPR/Cas9
    He Z, Proudfoot C, et al
    Biotechnol Bioeng, [Epub ahead of print]: doi: 10.1002/bit.25490 (2014)
    gBlocks Gene Fragments are used to generate sgRNA sequences for targeting Cas9
  10. Guide RNA Functional Modules Direct Cas9 Activity and Orthogonality
    Briner AE, Donohoue PD, et al
    Cell, 56(2):333–339 (2014)
    gBlocks Gene Fragments were used to create sgRNA expression plasmids.
  11. Programmable RNA recognition and cleavage by CRISPR/Cas9
    O'Connell MR, Oakes BL, et al
    Nature, 516(7530):263–266 (2014)
    gBlocks Gene Fragments were used to create sgRNA, gene specific sequences.
  12. A Homology Integrated CRISPR-Cas (HI-CRISPR) system for one-step multi-gene disruptions in Saccharomyces cerevisiae
    Bao Z, Xiao H, et al
    ACS Synth Biol, [Epub ahead of print]: (2014)
  13. Optimized CRISPR/Cas tools for efficient germline and somatic genome engineering in Drosophila
    Port F, Chen HM, et al
    Proc Natl Acad Sci U S A, 111(29):2967–76 (2014)
    gBlocks Gene Fragments were used to construct a codon optimized Cas9 sequence for expression in Drosophila. Optimization of the sequence was performed using the IDT Codon Optimization tool.
  14. CRISPR/Cas9-Directed Genome Editing of Cultured Cells
    Yang L, Yang JL, et al
    Curr Protoc Mol Biol, doi: 10.1002/0471142727:1–17 (2014)
    The authors provide a series of protocols for CRISPR/Cas9 genome editing in cell culture. The protocol describes how to use gBlocks Gene Fragments to generate the necessary guide RNAs and apply them to cell culture experiments.
  15. Genome editing in the human malaria parasite Plasmodium falciparum using the CRISPR-Cas9 system
    Ghorbal M, Gorman M, et al
    Nat Biotechnol, 32(8):819–821 (2014)
    A gBlocks Gene Fragment was used to create an sgRNA expression vector for use with the CRISPR/Cas9 system.
  16. Accelerating genome editing in CHO cells using CRISPR Cas9 and CRISPy, a web-based target finding tool
    Ronda C, Pedersen LE, et al
    Biotechnol Bioeng, 111(8):1604–1616 (2014)
    A gBlocks Gene Fragment was used to generate an sgRNA expression cassette for use in CRISPR/Cas9 genome editing experiments in CHO cells.
  17. Generation of human endometrial knockout cell lines with the CRISPR-Cas9 system confirms the prostaglandin F2α synthase activity of aldo-ketoreductase 1B1
    Lacroix Pépin N, Chapdelaine P, et al
    Mol Hum Reprod, 20(7):650–663 (2014)
    A gBlocks Gene Fragment was used to generate a single-guide RNA scaffold and cloned into a vector for use in CRISPR/Cas9 genome editing.
  18. Dynamic Imaging of Genomic Loci in Living Human Cells by an Optimized CRISPR/Cas System
    Chen B, Gilbert LA, et al
    Cell, 155:1479–1491 (2013)
    gBlocks Gene Fragments were used to generate expression constructs for optimized sgRNAs, that were then used to target a nuclease-deficient, Cas9-GFP chimeric protein in chromatin imaging experiments.
  19. Mutagenesis and homologous recombination in Drosophila cell lines using CRISPR/Cas9
    Bassett AR, Tibbit C, et al
    Biol Open, 3(1):42–49 (2013)
    A gBlocks Gene Fragment was used to generate a sgRNA backbone, including a Drosophila U6 promoter.
  20. Repurposing CRISPR/Cas9 for in situ functional assays
    Malina A, Mills JR, et al
    Genes & Dev, 27:2602–2614 (2013)
    gBlocks Gene Fragments were used to generate a sgRNA constructs for in situ functional assays.
  21. Orthogonal Cas9 proteins for RNA-guided gene regulation and editing
    Esvelt KM, Mali P, et al
    Nat Methods, 10(11):1116–1121 (2013)
    The authors of this study, use IDT gBlocks® Gene Fragments to generate human optimized Cas9 orthologs from  S. thermophilus, N. meningitidis, and T. denticola, as well as tracRNA expression cassettes for each ortholog. The authors clarify the PAM requirements of the various enzymes, and show that combining Cas9 orthologs with a different PAM requirements can be used to carry out different tasks in parallel in bacterial and human cell types.
  22. Engineering the Caenorhabditis elegans genome using Cas9-triggered homologous recombination
    Dickinson DJ, Ward JD, et al
    Nat Methods, 10(10):1028–1034 (2013)
    A codon-optimized Cas9 was created by ordering a series of overlapping gBlocks Gene Fragments, and assembled using the Gibson Assembly™ Method, and inserted into the vector pCFJ601. In addition, a single gBlocks Gene Fragment was inserted downstream of the Cas9 sequence, containing a U6 promoter, and the necessary elements for sgRNA function.
    CRISPR/Cas9, Gibson Assembly Method
  23. RNA-guided gene activation by CRISPR-Cas9-based transcription factors
    Perez-Pinera P, Kocak DD, et al
    Nat Methods, 10(10):973–976 (2013)
    gBlocks Gene Fragments were used to generate crRNA and tracRNA expression cassettes, as well as a single-guide RNA expression cassette that was designed for easy cloning of future RNA spacer elements.
  24. Genome Engineering of Drosophila with the CRISPR RNA-Guided Cas9 Nuclease
    Gratz SJ, Cummings AM, et al
    Genetics, 194(4):1029–1035 (2013)
    A gBlocks Gene Fragment was used to generate a chimeric single-guide RNA expression cassette with a U6 promoter that was designed for easy cloning of RNA spacer elements.
  25. CRISPR/Cas9-Targeted Mutagenesis in Caenorhabditis elegans
    Waaijers S, Portegijs V, et al
    Genetics, 195(3):1187–1191 (2013)
    gBlocks Gene Fragments containing either a T7 or U6 promoter and the necessary tracrRNA and crRNA elements were used to create expression vectors for single guide RNAs (sgRNA) for targeting Cas9 to desired targets in C. elegans.
  26. Multiplexed activation of endogenous genes by CRISPR-on, an RNA-guided transcriptional activator system
    Cheng AW, Wang H, et al
    Cel Res, 23(10):1163–1171 (2013)
    A gBlocks Gene Fragment was used to add 10 repeats of the minimal VP16 transactivation domain, derived from herpes simplex virus, to a nuclease deficient Cas9. This allows the resulting dCas9VP160 chimeric protein to serve as a transcriptional activator that can be easily target by CRISPR machinery in a variety of cell types.
  27. CAS9 transcriptional activators for target specificity screening and paired nickases for cooperative genome engineering
    Mali P, Aach J, et al
    Nat Biotech, advance online publication: (2013)

    gBlocks Gene Fragments were used for creating sgRNA expression constructs for targeting mutant Cas9 variants to test either transcription activation or genome modification.

  28. Heritable genome editing in C. elegans via a CRISPR-Cas9 system
    Friedland A, Tzur Y, et al
    Nat Methods, 10(8):741–743 (2013)
    gBlocks Gene Fragments were used to create an RNA Pol III expression vector that was then used to create single guide RNAs (sgRNAs) for Cas 9 target genes, unc-119, dpy-13, klp1, and Y61A9LA.1 in C. elegans.
  29. CRISPR-Mediated Modular RNA-Guided Regulation of Transcription in Eukaryotes
    Gilbert LA, Larson MH, et al
    Cell, 154(2):422–51 (2013)
    gBlocks Gene Fragments were used to create an Mxi1 transcription repressor domain and assembled with a dCas9 construct using the Gibson Assembly™ method into an expression vector. The resulting chimeric, transcriptional repressor was then shown to be targetable using the CRISPR/Cas9 mechanism.
    CRISPR/Cas9, Gibson Assembly Method
  30. RNA-guided human genome engineering via Cas9
    Mali P, Yang L, et. al
    Science, 336(6121):823–826 (2013)

    Use of gBlocks® Gene Fragments to assemble a sequence-optimized Cas9 for expression in human cells, as well as a Cas9_D10A variant.

    gBlocks, CRISPR/Cas9