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Real-time isothermal RNA amplification of toxic marine microalgae using preserved reagents on an integrated microfluidic platform

Real-time isothermal RNA amplification of toxic marine microalgae using preserved reagents on an integrated microfluidic platform
Real-time isothermal RNA amplification of toxic marine microalgae using preserved reagents on an integrated microfluidic platform
Quantitation of specific RNA sequences is a useful technique in marine biology that can elucidate cell abundance, speciation and viability, especially for early detection of harmful algal blooms. We are thus developing an integrated microfluidic system for cell concentration and lysis, RNA extraction/purification and quantitative RNA detection for environmental applications. The portable system is based on a microfluidic cartridge, or “lab-card”, using a low-cost injection moulded device, with a laminated lid. Here we present real-time isothermal RNA amplification using reagent master-mixes preserved on-chip in a gel at 4 °C for up to eight months. We demonstrate quantitation by reference to an internal control in a competitive assay with 500 cell equivalents of the toxic microalga Karenia brevis. Annealing of primers, amplification at 41 °C and real-time fluorescence detection of the internal control and target using sequence-specific molecular beacons were all performed on-chip
0003-2654
593-602
Tsaloglou, Maria-Nefeli
99ab30ba-15da-4d25-86ba-608d127f8369
Laouenan, Florian
37796527-85f1-42fd-a45d-c119bafe702d
Loukas, Christos-Moritz
cda31664-e0d8-46d9-b9d2-162484c18baf
Monsalve, Lisandro Gabriel
43f42898-6a3c-4a4e-9bf8-30a77e621932
Thanner, Christine
4700ae27-009a-4141-a15d-8f2a2d94ed5b
Morgan, Hywel
de00d59f-a5a2-48c4-a99a-1d5dd7854174
Ruano-López, Jesus M.
d4ff6945-0de2-42b7-a225-4fb68385a018
Mowlem, Matthew C.
6f633ca2-298f-48ee-a025-ce52dd62124f
Tsaloglou, Maria-Nefeli
99ab30ba-15da-4d25-86ba-608d127f8369
Laouenan, Florian
37796527-85f1-42fd-a45d-c119bafe702d
Loukas, Christos-Moritz
cda31664-e0d8-46d9-b9d2-162484c18baf
Monsalve, Lisandro Gabriel
43f42898-6a3c-4a4e-9bf8-30a77e621932
Thanner, Christine
4700ae27-009a-4141-a15d-8f2a2d94ed5b
Morgan, Hywel
de00d59f-a5a2-48c4-a99a-1d5dd7854174
Ruano-López, Jesus M.
d4ff6945-0de2-42b7-a225-4fb68385a018
Mowlem, Matthew C.
6f633ca2-298f-48ee-a025-ce52dd62124f

Tsaloglou, Maria-Nefeli, Laouenan, Florian, Loukas, Christos-Moritz, Monsalve, Lisandro Gabriel, Thanner, Christine, Morgan, Hywel, Ruano-López, Jesus M. and Mowlem, Matthew C. (2013) Real-time isothermal RNA amplification of toxic marine microalgae using preserved reagents on an integrated microfluidic platform. Analyst, 138 (2), 593-602. (doi:10.1039/c2an36464f).

Record type: Article

Abstract

Quantitation of specific RNA sequences is a useful technique in marine biology that can elucidate cell abundance, speciation and viability, especially for early detection of harmful algal blooms. We are thus developing an integrated microfluidic system for cell concentration and lysis, RNA extraction/purification and quantitative RNA detection for environmental applications. The portable system is based on a microfluidic cartridge, or “lab-card”, using a low-cost injection moulded device, with a laminated lid. Here we present real-time isothermal RNA amplification using reagent master-mixes preserved on-chip in a gel at 4 °C for up to eight months. We demonstrate quantitation by reference to an internal control in a competitive assay with 500 cell equivalents of the toxic microalga Karenia brevis. Annealing of primers, amplification at 41 °C and real-time fluorescence detection of the internal control and target using sequence-specific molecular beacons were all performed on-chip

Full text not available from this repository.

More information

Published date: 2013
Organisations: Electronics & Computer Science, Ocean Technology and Engineering

Identifiers

Local EPrints ID: 346731
URI: https://eprints.soton.ac.uk/id/eprint/346731
ISSN: 0003-2654
PURE UUID: 3603676e-7eca-4298-9924-07ecd39c4e26
ORCID for Hywel Morgan: ORCID iD orcid.org/0000-0003-4850-5676

Catalogue record

Date deposited: 08 Jan 2013 10:55
Last modified: 20 Jul 2019 00:59

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