General Notes

General Notes

Plants

Preparation of Plant Agar Media
Arabidopsis Seeding Protocol
Protocol for Removing Fungal Infections

Extraction

TRIzol Extraction Protocol
Whatman No. 1 Paper Extraction Protocol
FTA Extraction Protocol
Salt Extraction Protocol
Isopropanol Extraction Protocol
Lysis Protocol
Farmer Extraction Protocol

Hybridisation

APTMS Addition to Paper
Binding the DNA Probe to Paper
Blocking the Paper for Hybridisation
Sandwich Hybridisation Step
UV Spectrophotometer Melt Curves
Melt Curve Testing With SYBR Green

RCA

Padlock Probe Circularisation
Polyacrylamide Gel Electrophoresis (PAGE)
DNA Extraction
RCA Proof

BRET

Electrocompetent E. coli Transformation
Protein Expression
Protein Purification Attempt 1
Protein Purification Attempt 2
Nano-Glo® Luciferase Assay System
mNeonGreen Fluorescence

G-Quadruplex

Preparation fo G-Quadruplex Assay Buffer
ThT Dilution Series
G-Quadruplex in Solution Phase: Simultaneously vs After
G-Quadruplex in Solution Phase: Standard Curve
Paper Disc Production
G-Quadruplex on Paper Discs

RT-qPCR

Designing and Folding of Stem-Loop Primers for RT-qPCR
Designing qPCR Primers and Probes and Pre-Formulating Primer-Probe Mixes for RT-qPCR
Performing RT-qPCR

References

Extraction

  1. Thermo Fisher Scientific. (2012, June 14). PureLink® RNA Mini Kit: User guide (Publication No. MAN0000406, Rev. 3.00) [PDF]. https://documents.thermofisher.com/TFS-Assets/LSG/manuals/purelink_rna_mini_kit_man.pdf
  2. Zou, Y., Mason, M. G., Wang, Y., Wee, E., Turni, C., Blackall, P. J., Trau, M., & Botella, J. R. (2017). Nucleic acid purification from plants, animals and microbes in under 30 seconds. PLOS Biology, 15(11), e2003916. https://doi.org/10.1371/journal.pbio.2003916
  3. Leong, S. M., Tan, K. M.-L., Chua, H. W., Huang, M.-C., Cheong, W. C., Li, M.-H., Tucker, S., & Koay, E. S.-C. (2017). Paper-Based MicroRNA Expression Profiling from Plasma and Circulating Tumor Cells. Clinical Chemistry, 63(3), 731–741. https://doi.org/10.1373/clinchem.2016.264432
  4. Nilsen, T. W. (2012). Selective precipitation of large RNAs [Protocol]. Cold Spring Harbor Protocols. https://doi.org/10.1101/pdb.prot072322
  5. Qiagen. (n.d.). Isopropanol precipitation of DNA [Web page]. https://www.qiagen.com/us/knowledge-and-support/knowledge-hub/bench-guide/dna/handling-dna/isopropanol-precipitation-of-dna
  6. MP Biomedicals. (n.d.). FastPrep‑24 Classic user manual [PDF]. https://www.mpbio.com/media/document/file/manual/dest/f/a/s/t/p/FastPrep-24_Classic_User_Manual.pdf

Hybridisation

  1. Li, X., & Zhou, W. (Inventors). (2021). One‑step surface modification to graft DNA codes on paper and its bio‑applications (U.S. Patent No. 11,155,812 B2). U.S. Patent and Trademark Office. https://patents.google.com/patent/US11155812B2/en
  2. Beaumont, M., Bacher, M., Opietnik, M., Gindl-Altmutter, W., Potthast, A., & Rosenau, T. (2018). A General Aqueous Silanization Protocol to Introduce Vinyl, Mercapto or Azido Functionalities onto Cellulose Fibers and Nanocelluloses. Molecules, 23(6), 1427. https://doi.org/10.3390/molecules23061427
  3. Clancy, E., Burke, M., Vahid Arabkari, Barry, T., Kelly, H., Dwyer, R. M., Kerin, M. J., & Smith, T. J. (2017). Amplification-free detection of microRNAs via a rapid microarray-based sandwich assay. Analytical and Bioanalytical Chemistry, 409(14), 3497–3505. https://doi.org/10.1007/s00216-017-0298-6
  4. Yaichi Kawakatsu, Okada, R., Hara, M., Tsutsui, H., Yanagisawa, N., Tetsuya Higashiyama, Arima, A., Baba, Y., Ken-ichi Kurotani, & Michitaka Notaguchi. (2024). Microfluidic device for simple diagnosis of plant growth condition by detecting miRNAs from filtered plant extracts. Plant Phenomics. https://doi.org/10.34133/plantphenomics.0162
  5. Zhou, F., Noor, M., & Krull, U. (2015). A Paper-Based Sandwich Format Hybridization Assay for Unlabeled Nucleic Acid Detection Using Upconversion Nanoparticles as Energy Donors in Luminescence Resonance Energy Transfer. Nanomaterials, 5(4), 1556–1570. https://doi.org/10.3390/nano5041556
  6. Wang, J., Pan, X., & Liang, X. (2016). Assessment for Melting Temperature Measurement of Nucleic Acid by HRM. Journal of Analytical Methods in Chemistry, 2016. https://doi.org/10.1155/2016/5318935
  7. Liu, Y., Zhan, L., Qin, Z., Sackrison, J., & Bischof, J. C. (2021). Ultrasensitive and Highly Specific Lateral Flow Assays for Point-of-Care Diagnosis. ACS Nano, 15(3), 3593–3611. https://doi.org/10.1021/acsnano.0c10035
  8. Schroeder, S. J., & Turner, D. H. (2009). Optical Melting Measurements of Nucleic Acid Thermodynamics. Methods in Enzymology, 371–387. https://doi.org/10.1016/s0076-6879(09)68017-4

RCA

  1. New England Biolabs. (2025, January 10). Monarch Spin RNA Cleanup Kit protocol. https://www.neb.com/en-gb/protocols/2025/01/10/monarch-spin-rna-cleanup-kit-protocol
  2. NEBExpress® Ni Spin Column Reaction Protocol (NEB #S1427) | NEB. (n.d.). https://www.neb.com/en-gb/protocols/2019/08/28/nebexpress-ni-spin-column-reaction-protocol-neb-s1427
  3. Li, Y., Yang, P., Lei, N., Ma, Y., Ji, Y., Zhu, C., & Wu, Y. (2018). Assembly of DNA-Templated Bioluminescent Modules for Amplified Detection of Protein Biomarkers. Analytical Chemistry, 90(19), 11495–11502. https://doi.org/10.1021/acs.analchem.8b02734
  4. Li, Y., Zhou, L., Ni, W., Luo, Q., Zhu, C., & Wu, Y. (2019). Portable and Field-Ready Detection of Circulating MicroRNAs with Paper-Based Bioluminescent Sensing and Isothermal Amplification. Analytical Chemistry, 91(23), 14838–14841. https://doi.org/10.1021/acs.analchem.9b04422
  5. Thermo Fisher Scientific. (2012). Denaturing polyacrylamide/urea gel electrophoresis (Manual No. MAN0011970). https://assets.fishersci.com/TFS-Assets/LSG/manuals/MAN0011970_Denaturing_PolyacrylamideUrea_Gel_Electrophoresis_UG.pdf
  6. Lucigen (Cambio). (2019, July). CircLigase II ssDNA ligase: Protocol manual (MA298E‑CircLigase II ssDNA Ligase). https://www.cambio.co.uk/library/images/html_images/2022%20protocols/manual_CircLigase-II-ssDNA-Ligase.pdf
  7. Summer H, Grämer R, Dröge P. Denaturing urea polyacrylamide gel electrophoresis (Urea PAGE). J Vis Exp. 2009 Oct 29;(32):1485. doi: 10.3791/1485. PMID: 19865070; PMCID: PMC3329804.
  8. Renaud de la Faverie, A., Guédin, A., Bedrat, A., Yatsunyk, L. A., & Mergny, J. L. (2014). Thioflavin T as a fluorescence light-up probe for G4 formation. Nucleic acids research, 42(8), e65. https://doi.org/10.1093/nar/gku111

RT-qPCR

  1. Kramer, M. F. (2011). Stem‐Loop RT‐qPCR for miRNAs. Current Protocols in Molecular Biology, 95(1). https://doi.org/10.1002/0471142727.mb1510s95
  2. Varkonyi-Gasic, E., Wu, R., Wood, M., Walton, E. F., & Hellens, R. P. (2007). Protocol: a highly sensitive RT-PCR method for detection and quantification of microRNAs. Plant Methods, 3(1), 12. https://doi.org/10.1186/1746-4811-3-12