RECOMMENDED READING

Dharmacon publications

Our scientists take pride in the fact that their contributions to the scientificcommunity go beyond research and development of experimental reagents.
Below is a list of peer-reviewed publications from research performed at
Horizon Discovery (formerly Dharmacon) and external collaborations.

CRISPR-Cas9 genome engineering
1. Anderson E.M., S. McClelland, et al. Lactobacillus gasseri CRISPR-Cas9
characterization In Vitro reveals a flexible mode of protospacer-adjacentmotif recognition. PLoS One. 13, e0192181 (2018). doi: 10.1371/journal.
pone.0192181
2. Basila, M., M. L. Kelley, et al.. Minimal 2’-O-methyl phosphorothioate
linkage modification pattern of synthetic guide RNAs for increased
stability and efficient CRISPR-Cas9 gene editing avoiding cellular
toxicity. PLoS One. 12, e0188593 (2017). doi: 10.1371/journal.
pone.0188593
3. Strezoska, Ž., M. Perkett, et al. High-content analysis screening for cell
cycle regulators using arrayed synthetic crRNA libraries. J. Biotechnol.
251, 189-200 (2017). doi: 10.1016/j.jbiotec.2017.04.017
4. He, K., E. Chou, et al. Conjugation and evaluation of triazole-linked single
guide RNA for CRISPR-Cas9 gene editing. ChemBioChem.
DOI: 10.1002/cbic.201600320 (2016).
5. Kelley, M.L., Ž. Strezoska, et al. Versatility of chemically synthesized guide
RNAs for CRISPR-Cas9 genome editing. J. Biotechnol. 233, 74-83 (2016).
6. Anderson, E.M., A. Haupt, et al. Systematic analysis of CRISPR-Cas9
mismatch tolerance reveals low levels of off-target activity. J. Biotechnol.
211, 56-65 (2015).
7. Barrangou, R., A. Birmingham, et al. Advances in CRISPR-Cas9 genome
engineering: lessons learned from RNA interference. Nucleic Acids Res.
43, 3407–3419 (2015).
8. Glemzaite, M., E. Balciunaite, et al. Targeted gene editing by transfectionof in vitro constituted Streptococcus thermophilus Cas9 nuclease
complex. RNA Biology. 12, 1–4 (2015).

RNAi library screening
1. Stombaugh, J., A. Licon, et al. The Power Decoder Simulator for the
Evaluation of Pooled shRNA Screen Performance. J. Biomolecular
Screening. 20, 965-75 (2015).
2. Freeley, M., E. Derrick, et al. RNAi Screening with Self-Delivering,
Synthetic siRNAs for Identification of Genes That Regulate Primary
Human T Cell Migration. J. Biomolecular Screening. 209, 943-956 (2015).

3. Vermeulen, A., A. van Brabant Smith, et al. Pooled shRNA Screening.
Frontiers in RNAi. 1, 58-78 (2014).
4. Couts, K.L., E.M. Anderson, et al. Oncogenic B-Raf signaling in melanoma
cells controls a network of microRNAs with combinatorial functions.
Oncogene. 32, 1959-1970 (2013).
5. Meliopoulos, V.A., L.E. Andersen, et al. Host gene targets for novel
influenza therapies elucidated by high-throughput RNA interference
screens. The FASEB Journal. 26, 1372-1386 (2012).
6. Schoolmeesters, A., D.D. Brown, et al. Kinome-Wide Functional
Genomics Screen Reveals a Novel Mechanism of TNFα-Induced Nuclear
Accumulation of the HIF-1α Transcription Factor in Cancer Cells. PloS
One. 7, e31270 (2012).
7. Strezoska, Ž., A. Licon, et al. Optimized PCR Conditions and Increased
shRNA Fold Representation Improve Reproducibility of Pooled shRNA
Screens. PloS One. 7, e42341 (2012).
8. Santhakumar, D., T. Forster, et al. Combined agonist-antagonist genomewide functional screening identifies broadly active antiviral microRNAs.
P. Natl. Acad. Sci-Biol. 107, 13830-13835 (2010).
9. Zumbansen, M., L.M. Altrogge, et al. First siRNA library screening in
hard-to-transfect HUVEC cells. Journal of RNAi and Gene Silencing: anInternational Journal of RNA and Gene Targeting Research. 6, 354 (2010).
10. Kwan, P., A. Birmingham, et al. NoiseMaker: simulated screens forstatistical assessment. Bioinformatics. 26, 2484-2485 (2010).
11. Birmingham, A., L.M. Selfors, et al. Statistical methods for analysis ofhigh-throughput RNA interference screens. Nat. Methods. 6, 569-575(2009).
12. K.J. Simpson, L.M. Selfors, et al. Identification of genes that regulate
epithelial cell migration using an siRNA screening approach. Nat. Cell Bio.
10, 1027-1038 (2008).
13. Kolokoltsov, A.A., D. Deniger, et al. Small interfering RNA profilingreveals key role of clathrin-mediated endocytosis and early endosome
formation for infection by respiratory syncytial virus. J. Virol. 81, 77867800 (2007).
14. Boese, Q.F., D. Samarsky, et al. siARRAY reverse transfection format (RTF):
a rapid method for RNAi-based high-throughput studies of biologicalpathways. Nat. Methods Application Notes. 1, (2006).
15. Hsieh, A.C., R. Bo, et al. A library of siRNA duplexes targeting thephosphoinositide 3-kinase pathway: determinants of gene silencing foruse in cell-based screens. Nucleic Acids Res. 32, 893-901 (2004).

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RNAi in vivo
1. Fernanda Lara, M., E. González-González, et al. Inhibition of CD44 Gene
Expression in Human Skin Models Using Self-Delivery Short Interfering
RNA Administered by Dissolvable Microneedle Arrays. Hum. Gene Ther.
23, 1-8 (2012).
2. Lobovkina, T., G.B. Jacobson, et al. In vivo sustained release of siRNAfrom solid lipid nanoparticles. ACS Nano. 5, 9977-9983 (2011).
3. Hickerson, R.P., M.A. Flores, et al. Use of self-delivery siRNAs to inhibit
gene expression in an organotypic pachyonychia congenita model.
J. Invest. Dermatol. 131, 1037-1044 (2011).
4. Jacobson, G.B., E. Gonzalez-Gonzalez, et al. Biodegradable nanoparticles
with sustained release of functional siRNA in skin. J. Pharm. Sci. 99, 42614266 (2010).
5. Ra, H., W. Piyawattanametha, et al. In vivo imaging of human and mouseskin with a handheld dual-axis confocal fluorescence microscope. J.
Invest. Dermatol. 131, 1061-1066 (2010).
6. Gonzalez-Gonzalez, E., T.J. Speaker, et al. Silencing of reporter gene
expression in skin using siRNAs and expression of plasmid DNA
delivered by a soluble protrusion array device (PAD). Mol. Ther. 18, 16671674 (2010).
7. Wang, Q., H. Ilves, et al. Delivery and inhibition of reporter genes bysmall interfering RNAs in a mouse skin model. J. Invest. Dermatol. 127,2577-2584 (2007).
8. Smith, F.J.D., R.P. Hickerson, et al. Development of therapeutic siRNAs forpachyonychia congenita. J. Invest. Dermatol. 128, 50-58 (2007).

RNAi
1. Mutisya, D., T. Hardcastle, et al. Amide linkages mimic phosphates in
RNA interactions with proteins and are well tolerated in the guide strandof short interfering RNAs. Nucleic Acids Res. (2017) gkx558. doi: 10.1093/
nar/gkx558
2. Mutisya, D., T. Hardcastle, et al. Amide linkages mimic phosphates in RNA
interactions with proteins and are well tolerated in the guide strand of
short interfering RNAs. Nucleic Acids Res. In press (2017).
3. Caoa, Y-A., R.P. Hickerson, et al. Gene expression profiling inpachyonychia congenita skin. J. Dermatol. Sci. 77, 156-165 (2015).
4. Cuellar, T.L., D. Barnes, et al. Systematic evaluation of antibody-mediatedsiRNA delivery using an industrial platform of THIOMAB-siRNA
conjugates. Nucleic Acids Res. 43, 1189-11203 (2015).
5. Birmingham, A., A. Kaufmann and K. Kozak. RNAi and Off-Target Effects.
Frontiers in RNAi. 1, 3-20 (2014).
6. Schmidt, E.E., M.S. Banos, et al. Public Repositories for RNAi Screening
Data. Frontiers in RNAi, 1, 40-57 (2014).
7. Hickerson, R.P., S.A. Leachman, et al. Development of quantitative
molecular clinical end points for siRNA clinical trials. J. Invest. Dermatol.
131, 1029-1036 (2010).
8. DiFeo, A., F. Huang, et al. KLF6-SV1 is a novel antiapoptotic proteinthat targets the BH3-only protein NOXA for degradation and whose
inhibition extends survival in an ovarian cancer model. Cancer Res. 69,
4733-4741 (2009).
9. Hickerson, R.P., D. Leake, et al. Rapamycin selectively inhibits expressionof an inducible keratin (K6a) in human keratinocytes and improves
symptoms in pachyonychia congenita patients. J. Dermatol. Sci. 56, 8288 (2009).
10. Baskin, L., S. Urschel and B. Eiberger. A novel ex-vivo application of RNAi
for neuroscience. Biotechniques. 45, 338-339 (2008).

11. Rugg, E.L., F.J.D. Smith, et al. Development of Therapeutic siRNAs for
Pachyonychia Congenita. Commentary. J. Invest. Dermatol. 128, (2008).
12. Hickerson, R.P., A.V. Vlassov, et al. Stability study of unmodified siRNAand relevance to clinical use. Oligonucleotides. 18, 345-354 (2008).
13. Hickerson, R.P., F.J.D. Smith, et al. Single-nucleotide-specific siRNAtargeting in a dominant-negative skin model. J. Invest. Dermatol.
128, 594-605 (2007).
14. Anderson, J.S., A. Vermeulen, et al. Complete Suppression of CCR5
Expression and Inhibition of HIV-1 Infection by Transfected andLentiviral Vector Expressed shRNAs. Mol. Ther. 13, S310 (2006).
15. Radisky, D.C., D.D. Levy, et al. Rac1b and reactive oxygen species mediate
MMP-3-induced EMT and genomic instability. Nature. 436, 123-127 (2005).
16. Anderson, E.M., P. Miller, et al. Gene profiling study of G3139-and Bcl-2targeting siRNAs identifies a unique G3139 molecular signature. Cancer
Gene Ther. 13, 406-414 (2005).
17. Benimetskaya, L., J.C. Lai, et al. Induction of apoptosis by G3139 inmelanoma cells. Ann. NY Acad. Sci. 1058, 235-245 (2005).
18. Lai, J.C., L. Benimetskaya, et al. Phosphorothioate oligodeoxynucleotides
and G3139 induce apoptosis in 518A2 melanoma cells. Mol. Cancer Ther.
4, 305-315 (2005).
19. Benimetskaya, L., J.C. Lai, et al. Relative Bcl-2 independence of druginduced cytotoxicity and resistance in 518A2 melanoma cells. ClinicalCancer Res. 10, 8371-8379 (2004).
20. Raffo, A., J.C. Lai, et al. Antisense RNA down-regulation of bcl-2
expression in DU145 prostate cancer cells does not diminish thecytostatic effects of G3139 (Oblimersen). Clinical Cancer Res. 10, 31953206 (2004).
21. Huang, F., A. Khvorova, et al. Analysis of clathrin-mediated endocytosis
of epidermal growth factor receptor by RNA interference. J. Biol. Chem.
279, 16657-16661 (2004).
22. Li, T., C-Y. Chang, et al. Identification of the gene for vitamin K epoxide
reductase. Nature. 427, 541-544 (2004).
23. Jiang, Z.Y., Q.L. Zhou, et al. Insulin signaling through Akt/protein kinase
B analyzed by small interfering RNA-mediated gene silencing. P. Natl.
Acad. Sci-Biol. 100, 7569-7574 (2003).

microRNA/noncoding RNA
1. Hodzic J., Sie Daoud, A. Vermeulen Annaleen, et al. Functional Screening
Identifies Human miRNAs that Modulate Adenovirus Propagation in
Prostate Cancer Cells. Human Gene Therapy. January 2017, ahead of print.
DOI: 10.1089/hum.2016.143.
2. Zhang, Z., J.E. Lee, et al. High-efficiency RNA cloning enables accuratequantification of miRNA expression by deep sequencing. Genome
Biology. 14, R109 (2013).
3. Robertson, B., A.B. Dalby, et al. Specificity and functionality of microRNA
inhibitors. Silence. 1, 2-9 (2010).
4. Schoolmeesters, A., T. Eklund, et al. Functional profiling reveals criticalrole for miRNA in differentiation of human mesenchymal stem cells.
PLoS One. 4, e5605 (2009).

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RNAi specificity & functionality

RNA/structure/synthesis

1. Brown, J.W., A. Birmingham, et al. The RNA structure alignment ontology.
RNA. 15, 1623-1631 (2009).
2. de Zwart I., et al. in Handbook of RNA Biochemistry, R. K. Hartmann, A.
Bindereif, Eds. (Wiley, New York, 2008) pp. 783-806.
3. Anderson, E.M., A. Birmingham, et al. Experimental validation of theimportance of seed complement frequency to siRNA specificity. RNA. 14,
853-861 (2008).
4. Anderson, E.M., Q.F. Boese, et al. Identifying siRNA-induced off-targets
by microarray analysis. Methods Mol. Bio. 442, 45 (2008).
5. Birmingham, A., E.M. Anderson, et al. A protocol for designing siRNAs
with high functionality and specificity. Nat. Protoc. 2, 2068-2078 (2007).
6. Vermeulen, A., B. Robertson, et al. Double-stranded regions are essential
design components of potent inhibitors of RISC function. RNA. 13, 723730 (2007).
7. Li, L., X. Lin, et al. Defining the optimal parameters for hairpin-based
knockdown constructs. RNA. 13, 1765-1774 (2007).
8. Birmingham, A., E.M. Anderson, et al. 3' UTR seed matches but not overall
identity are associated with RNAi off-targets. Nat. Methods. 3, 199-204 (2006).
9. Reynolds, A., E.M. Anderson, et al. Induction of the interferon response by
siRNA is cell type- and duplex length-dependent. RNA. 12, 988-993 (2006).
10. Fedorov, Y., E.M. Anderson, et al. Off-target effects by siRNA can induce
toxic phenotype. RNA. 12, 1188-1196 (2006).
11. Jackson, A.L., J. Burchard, et al. Position-specific chemical modification ofsiRNAs reduces “off-target” transcript silencing. RNA. 12, 1197-1205 (2006).
12. Boese, Q.F., W.S. Marshall, et al. Design and synthesis of small interfering
RNA (siRNA) in RNA Interference Technology: From Basic Science to Drug
Development, (Cambridge Univ. Press, 2005), chap. 6, pp. 103-117.
13. Khvorova, A., Boese, Q. and Marshall, W. S. (2005) Rational siRNA
Design for RNA Interference: Optimizations for Therapeutic Use
and Current Applications, in Modern Biopharmaceuticals: Design,
Development and Optimization (ed J. Knäblein), Wiley-VCH Verlag
GmbH, Weinheim, Germany.
14. Fedorov, Y., A. King, et al. Different delivery methods—different
expression profiles. Nat. Methods. 2, 241-241 (2005).
15. Boese, Q.F., D. Leake, et al. Mechanistic insights aid computational shortinterfering RNA design. Method Enzymol. 392, 73-96 (2005).
16. Vermeulen, A., L. Behlen, et al. The contributions of dsRNA structure to
Dicer specificity and efficiency. RNA. 11, 674-682 (2005).
17. Reynolds, A., D. Leake, et al. Rational siRNA design for RNA interference.
Nat. Biotechnol. 22, 326-330 (2004).
18. Karpilow, J., D. Leake, et al. siRNA: enhanced functionality through rational
design and chemical modification. PharmaGenomics. 32, 40 (2004).
19. Jackson, A.L., S.R. Bartz, et al. Expression Profiling reveals off-target gene
regulation by RNAi. Nat. Biotechnol. 21, 635-637 (2003).
20. Boese, Q.F., S.A. Scaringe, et al. siRNA as a tool for streamlining functional
genomic studies. Targets. 2, 93-100 (2003).
21. Khvorova, A., A. Reynolds, et al. Functional siRNAs and miRNAs exhibit
strand bias. Cell. 115, 209-216 (2003).

1. Wang, R., Z. Lou, et al. Base pairing and structural insights into the
5-formylcytosine in RNA duplex. Nucleic Acids Res. 44, 4968-4977 (2016).
2. Tanui, P., S.D. Kennedy, et al. Synthesis, biophysical studies and RNAinterference activity of RNA having three consecutive amide linkages.
Org. Biomol. Chem. 12, 1207-1210 (2014).
3. Mutisya, D., C. Selvam, et al. Amides are excellent mimics of phosphateinternucleoside linkages and are well tolerated in short interfering RNAs.
Nucleic Acids Res. 42, 6542-51 (2014).
4. Cantara, W.A., Y. Bilbille, et al. Modifications modulate anticodon loop
dynamics and codon recognition of E. coli tRNAArg1, 2. J. Mol. Bio.
416, 579-597 (2012).
5. Bilbille, Y., E.M. Gustilo, et al. The Human Mitochondrial tRNAMet:
Structure/Function Relationship of a Unique Modification in the
Decoding of Unconventional Codons. J. Mol. Bio. 406, 257-274 (2011).
6. Lusic, H., E.M. Gustilo, et al. Synthesis and investigation of the
5-formylcytidine modified, anticodon stem and loop of the humanmitochondrial tRNAMet. Nucleic Acids Res. 36, 6548-6557 (2008).
7. Delaney, M.O., A. Thomas, et al. Chromophoric 5'-O-Silyl Protection of
N-Protected 2'-ACE Ribonucleosides for Solid-Phase RNA Synthesis. Curr.
Protoc. Nucleic Acid Chem. 2.14, 2-14 (2008).
8. Scaringe, S.A., D. Kitchen, et al. Preparation of 5’-Silyl-2'Orthoester
Ribonucleosides for Use in Oligoribonucleotide Synthesis. Curr. Protoc.
Nucleic Acid Chem. 2-10 (2004).
9. Marshall, W.S., R.J. Kaiser, et al. Recent advances in the high-speed solid
phase synthesis of RNA. Curr. Opin. Chem. Bio. 8, 222-229 (2004).

Protein coding
1. Wiemann, S., C. Pennacchio, et al. The ORFeome Collaboration: agenome-scale human ORF-clone resource. Nature Methods. 13,
191–192 (2016).

Detection/qPCR/RT-qPCR
2. Strezoska, Ž., Y. Fedorov, et al. hMSC differentiation marker detection
using Thermo Scientific Solaris™ qPCR Gene Expression Assays. Nat.
Methods Application Notes. 6, (2009).

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©2018 Horizon Discovery Group Company—All rights reserved. First published December 2017. UK Registered Head Office: Building 8100, Cambridge
Research Park, Cambridge, CB25 9TL, United Kingdom.

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