A workflow for the relative quantification of multiple fish species from oceanic water samples using environmental DNA (eDNA) to support large-scale fishery surveys

A workflow for the relative quantification of multiple fish species from oceanic water samples using environmental DNA (eDNA) to support large-scale fishery surveys post thumbnail image
Whereas the variety of revealed marine research utilizing environmental DNA (eDNA) has elevated considerably in recent times, marine fish surveys are nonetheless scarce. To look at the potential for eDNA to assist marine fisheries monitoring surveys, we optimized an eDNA isolation technique, developed a multispecies assay and examined it on eDNA samples collected alongside the Pacific coast of the US.
4 business DNA extraction kits that exploit the potential of the nucleic acids binding a strong section (two utilizing a silica matrix and two magnetic beads) as effectively an natural separation technique had been examined. A species-specific multiplex qPCR assay was developed and examined to concurrently goal Pacific hake (Merluccius productus), Pacific lamprey (Entosphenus tridentatus) and eulachon (Thaleichthys pacificus).
The specificity of the assay was examined in silico, in vitro and in natura. Environmental DNA isolation utilizing phenol:chloroform:isoamyl purification with a section lock was optimized and yielded the best quantity of whole and goal DNA and was used to extract 46 marine water samples for the detection of the three species of curiosity.
The multiplex qPCR assay used within the quantification course of was additionally optimized to offer comfort and accuracy. Pacific hake was current in 44% of the eDNA samples whereas the opposite two species had been absent. Right here, we current an entire workflow for the simultaneous detection and quantification of a number of marine fish species utilizing eDNA.
This workflow helps large-scale at-sea sampling efforts with preservation at ambient temperatures and has demonstrated DNA extraction effectivity and reliability. The multiplex qPCR assay is proven to be delicate and particular for the needs of concurrently monitoring the relative abundance of a number of focused fish species.

Excessive salt transcription of DNA co-tethered with T7 RNA polymerase to beads generates elevated yields of extremely pure RNA

Excessive yields of RNA are routinely ready following the two-step strategy of high-yield in vitro transcription utilizing T7 RNA polymerase adopted by intensive purification utilizing gel separation or chromatographic strategies. We not too long ago demonstrated that in high-yield transcription reactions, as RNA accumulates in resolution, T7 RNA polymerase rebinds and extends the encoded RNA (utilizing the RNA as a template), leading to a product pool contaminated with longer-than-desired, (partially) double-stranded impurities.
Present purification strategies usually fail to completely remove these impurities which, if current in therapeutics, can stimulate the innate immune response with doubtlessly deadly penalties. On this work, we introduce a novel in vitro transcription technique that generates excessive yields of encoded RNA with out double-stranded impurities, lowering the necessity for additional purification.
Transcription is carried out at excessive salt circumstances to remove RNA product rebinding, whereas promoter DNA and T7 RNA polymerase are co-tethered in shut proximity on magnetic beads to drive promoter binding and transcription initiation, leading to a rise in total yield and purity of solely the encoded RNA.
A extra full elimination of double-stranded RNA throughout synthesis is not going to solely scale back total manufacturing prices, but in addition ought to finally allow therapies and applied sciences which can be at present being hampered by these impurities.

A conveyable magnetofluidic platform for detecting sexually transmitted infections and antimicrobial susceptibility

Efficient therapy of sexually transmitted infections (STIs) is proscribed by diagnostics that can’t ship outcomes quickly whereas the affected person continues to be within the clinic. The gold normal strategies for identification of STIs are nucleic acid amplification checks (NAATs), that are too costly for widespread use and have prolonged turnaround instances.
To handle the necessity for quick and reasonably priced diagnostics, we’ve got developed a transportable, speedy, on-cartridge magnetofluidic purification and testing (PROMPT) polymerase chain response (PCR) check. We present that it may detect Neisseria gonorrhoeae, the pathogen inflicting gonorrhea, with simultaneous genotyping of the pathogen for resistance to the antimicrobial drug ciprofloxacin in <15 min.
The duplex check was built-in right into a low-cost thermoplastic cartridge with automated processing of penile swab samples from sufferers utilizing magnetic beads. A compact instrument carried out DNA extraction, PCR, and evaluation of outcomes whereas relaying information to the person through a smartphone app.
This platform was examined on penile swab samples from sexual well being clinics in Baltimore, MD, USA (n = 66) and Kampala, Uganda (n = 151) with an total sensitivity and specificity of 97.7% and 97.6%, respectively, for N. gonorrhoeae detection and 100% concordance with tradition outcomes for ciprofloxacin resistance.
This examine paves the way in which for delivering accessible PCR diagnostics for quickly detecting STIs on the level of care, serving to to information therapy choices and fight the rise of antimicrobial resistant pathogens.

Isolation of goal DNA utilizing synergistic magnetic bead transport and electrokinetic movement

The arrival and dissemination of next-generation sequencing (NGS) applied sciences corresponding to Illumina’s sequencing platforms has introduced forth huge reductions in the fee, time, and technical difficulties related to DNA and RNA sequencing. Regardless of this development, the workflow required to generate nucleic acid libraries for sequencing stays time-consuming and laborious.
The next analysis proposes a way for simplifying and streamlining this course of by changing the guide washing steps of the widespread magnetic bead-based cleanup with a novel microfluidic technique by integrating magnetic separation and electrokinetic purification (MSEP).
Requiring no pumps, pipette mixing, vortexing, or centrifugation, MSEP depends on selective adsorption of goal DNA onto the magnetic beads with subsequent transport of beads by a microchannel present process an antiparallel electroosmotic movement. The synergetic movement circumstances had been optimized utilizing a easy electrohydrodynamic movement mannequin.
This work demonstrates that MSEP is as efficient in eliminating adapter-dimers from the post-ligation library combine because the guide technique whereas additionally tremendously lowering the hands-on time and quantity of pipetting required. Though MSEP has been utilized particularly towards NGS library preparation at the moment, it has the potential to be tailored and employed for any bead-based separation scheme, particularly, strong section extraction, sequence-specific hybridization, and immunoprecipitation on a microscale.
Greater-order superstructures of particular person DNA origami constructing blocks are incessantly utilized in DNA nanotechnology so as to improve the construction dimensions and complexity. Right here, a purification technique is offered to particularly enrich a totally assembled superstructure out of an extra of substructures.

Stool DNA Purification Kit

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Saliva DNA Purification Kit

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Saliva DNA Purification Kit

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EpiNext DNA Purification HT System

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  • EUR 234.30
  • EUR 1103.30
  • EUR 5052.30
  • 480 ml
  • 4 ml
  • 8 ml
  • 64 ml
  • 480 ml

Zenoprep Serum DNA Purification Kit

Z2005-000 Adjustable
EUR 574.8

Mini Spincolumn for DNA Purification

S1910-250 250 column
EUR 213.6

Mini Spincolumn for DNA Purification

S1910-500 500 column
EUR 310.8

Mini Spincolumn for DNA Purification

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Zenoquick Viral DNA Purification Kit

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Zenoquick Viral DNA Purification Kit

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Zenoprep Genomic DNA Purification Kit

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Zenoquick Genomic DNA Purification Kit

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Hair, Nails & Feathers DNA Purification Kit

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Hair, Nails & Feathers DNA Purification Kit

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Genomic DNA Extraction and Purification

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Plasmid DNA Extraction and Purification

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T-Pro Gel/PCR DNA Purification Maxi Kit (10)

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Agarose Gel DNA Extraction and Purification

EP10010 100 Tests
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The strategy relies on pull-down reactions with magnetic beads, the place superstructures are captured through an anchor strand on a selected terminus after which turn out to be separated from terminus-free constructions. By finishing up a number of pull-down reactions sequentially on completely different termini, the total superstructures that possess all termini turn out to be lastly enriched.
The strategy is demonstrated by purifying linear origami superstructures with as much as 9 monomers by two-sided pull-down reactions and a T-shaped superstructure in a three-sided pull-down response.

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