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A faster, smaller, better way to qPCR
Features & Benefits
Ultra-Fast Data Acquisition

35 cycles in 25 minutes

Unrivaled Performance

Detect 2-fold expression level differences

Portable & Compact

4.5 lbs – transport without ever calibrating

Scalable & Wireless

Connect up to 10 instruments
(48 samples/instrument)

Magnetic Induction Technology

Eliminate variability vs block-based cyclers

 

Q is intended for molecular biology applications. This product is not intended for the diagnosis, prevention or treatment of a disease.

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Q – qPCR Instrument

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Kit Size:  Q 2-channel qPCR Instrument
Part Number:  95900-2C
Price:  $12,700.00
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Kit Size:  Q 4-channel qPCR Instrument
Part Number:  95900-4C
Price:  $15,440.00
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HRM License for Q qPCR Instrument

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Kit Size:  1 HRM License
Part Number:  95915-20
Price:  $2,107.00
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Q Extended Warranty SILVER

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Kit Size:  Q Extended Warranty SILVER
Part Number:  95900-WSIL
Price:  $4,500.00
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Q Extended Warranty GOLD

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Kit Size:  Q Extended Warranty GOLD
Part Number:  95900-WGOL
Price:  $6,500.00
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Q Tubes & Caps (20 racks/box)

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Kit Size:  Q Tubes & Caps (20 racks/box)
Part Number:  95910-20
Price:  $133.00
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Description

Q uses a patented magnetic induction technology to rapidly heat samples coupled with fan forced air for cooling to acquire data in only 25 minutes. Available as a 4 channel model, the robust optical system acquires all channels simultaneously and allows for running the fastest multiplexed assays.

Q's miniature speaker-size and 4.5 pound weight make it the most portable and versatile qPCR machine on the market without ever needing to calibrate. Q also provides scalability as each qPCR machine can process up to 48 samples per run and up to 10 Q's can be connected to a single computer wirelessly via bluetooth enabling up to 480 samples to be processed simultaneously.

A key difference is that Q incorporates a unique spinning aluminum rotor providing superior temperature uniformity of ± 0.05°C versus traditional block-based real time thermal cyclers which rely on multiple peltier heating blocks that can create edge effects resulting in sample variation. Not only does the data give you superior reproducibility, repeatability but enables detection of 2-fold gene expression level differences as well as identification of difficult class IV SNP's requiring melt temperature resolutions of 0.1°C.

Who wouldn't want to take one for a spin?

 

See our Warranty Policy

 

Request a Demo

Register / Download Software

 


 

Meet the Quantabio Q cycler

 


 

Q-qPCR Software Tutorial – Overview

Q-qPCR Software Tutorial – Creating an Assay

Q-qPCR Software Tutorial – Starting a Run

 


 

Customer Profile Story
Developing a Fast and Sensitive qPCR Assay to Improve Field-Based TB Testing in Africa

Customer Profile Story

 

Customer Profile Story
Brewers Craft Disruptive Quality Standard Using the Latest Molecular Technologies

Customer Profile Story

 


 

Q Redefines Run Times to 25 Minutes

Details

Details

Contents

Specifications

Front Back

Physical

Height 5.1 in
Width 5.9 in
Length 5.9 in
Weight 4.5 lbs

 

Mic qPCR Cycler Specifications Front View     Mic qPCR Cycler Specifications Top View 

 


 

Thermal Performance

Temperature Accuracy ±0.25°C
Temperature Uniformity ±0.05°C
Ramp Rates Heating 4°C/s
Cooling 3°C/s
Temperature Input Range 40 – 99°C

 


 

Optical

Detectors Photodiode per channel
Excitation Sources High power LED for each channel
Channels Green Ex 465nm Em 510nm
Yellow Ex 540nm Em 570nm
Orange Ex 585nm Em 618nm
Red Ex 635nm Em 675nm
Acquisition Time 1 second

 


 

Reaction Tubes

Samples per Instrument 48
Reaction Volume Range 5 – 30µL

 


 

Operating Environment

Temperature 18 – 35ºC
Relative Humidity 20 – 80%

 

Customer Testimonials

Customer Testimonials

Q qPCR Instrument
My group was looking for a qPCR machine that didn’t take up much bench space or require annual maintenance, which the Q machine fits. Additional features I like include its ability to multiplex with up to 4 dyes in the same tube, its relative ease of use, and free software that can be used on anyone’s PC making it easy for everyone in the group to use. We’ve just begun using the machine, but so far I’ve been happy with the data acquisition obtained from it.
Max Rogers | Associate Director, Pathology and Toxicology Sciences
Q RT-PCR machine in Mbabane Swaziland
We had the pleasure of testing out the ToughMix Mastermix and the “Q” RT-PCR machine in Mbabane Swaziland in our Tuberculosis Centre of Excellence.” As it’s name implies, the ToughMix stood up to its name and made the 38 hour trip from Houston to Mbabane, Swaziland and had beautiful Ct curves. Similarly, the Q PCR machine was an ease to travel with as carry on luggage. It’s 25 minute run gave Ct curves identical to our existing, and much bulkier, RT-PCR machine that was already on site in Swaziland. Considering it’s size, intuitive software, and near perfect reproducible Ct curves, we’ve decided to add the Q as our workhorse RT-PCR machine
Assistant Professor | Infectious Diseases Global and Immigrant Health Baylor College of Medicine
Q qPCR Instrument

“We had the pleasure of testing the ToughMix and the “Q” RT-PCR machine in Mbabane Swaziland in our Tuberculosis Centre of Excellence. The ToughMix stood up to it’s name and had beautiful Ct’s….Considering it’s size, intuititive software and near reproducible Ct curves we’ve decided to add the Q as our workhorse.”

Assistant Professor | Infectious Diseases Global and Immigrant Health, Baylor College of Medicine
LAMP Assay on Q

"Performing a LAMP assay using a Q-qPCR machine is really comparable with Genie II, however, the higher number of samples that can be analysed at a time and also the appearance of the first amplification signals 2 min earlier than those in Genie II, adds up to its value!"

Steve Baeyen | Flanders Research Institute for agriculture, fisheries and food

Details

Contents

Specifications

Front Back

Physical

Height 5.1 in
Width 5.9 in
Length 5.9 in
Weight 4.5 lbs

 

Mic qPCR Cycler Specifications Front View     Mic qPCR Cycler Specifications Top View 

 


 

Thermal Performance

Temperature Accuracy ±0.25°C
Temperature Uniformity ±0.05°C
Ramp Rates Heating 4°C/s
Cooling 3°C/s
Temperature Input Range 40 – 99°C

 


 

Optical

Detectors Photodiode per channel
Excitation Sources High power LED for each channel
Channels Green Ex 465nm Em 510nm
Yellow Ex 540nm Em 570nm
Orange Ex 585nm Em 618nm
Red Ex 635nm Em 675nm
Acquisition Time 1 second

 


 

Reaction Tubes

Samples per Instrument 48
Reaction Volume Range 5 – 30µL

 


 

Operating Environment

Temperature 18 – 35ºC
Relative Humidity 20 – 80%

 

Performance Data

Resources

FAQs

  • Why are some Taq suited for fast cycling and others require the standard cycling profile?
    There are several commercially available Taq polymerases that differ in their enzymatic properties. Some display high processivity at temperatures above 60oC and are labelled as ‘fast cycling compatible’ polymerases because they can achieve PCR with a relatively short single annealing/extension step (<20 sec) without the need for a dedicated extension step at 68-72oC. These fast cycling compatible enzymes should be used with the ‘fast taq’ profile in the Q-qPCR software for details on how to set up a thermal profile. For Taq polymerases that are not compatible with fast cycling conditions, some consideration must be taken to select the appropriate thermal profile. For assays with an annealing temperature <60oC, a two-step cycling may not be achievable and a dedicated extension step and 68-72oC may be required. For assays with an annealing temperature >60oC, it may be possible to achieve a two-step cycling if the ‘standard taq’ profile is selected. This is because the standard taq setting will decrease the speed of transition between annealing and melt temperatures, which will increase the time that the reaction is held in the range of taq activity (a fast transition from annealing to melt temperature will quickly pass through this temperature range, while slowing this rate of increase will provide more time for taq to achieve amplicon extension).
  • What is the ramp rate?
    We do not typically like to discuss ramp rates, except to point out that they are an arbitrary measurement that may be misleading. A ramp rate is the rate of temperature change over time, but there is no standardisation for how to take this measurement. For instance, you could measure a ramp rate for a metal component of a machine that in reality has no relationship with the heat over a block, and will certainly not reflect your sample reactions temperature. The lack of similarity in ways to define and/or measure ramp rates makes the calculated values not comparable (it is like comparing how much you like something. One person likes it heaps, another lots. Its meaningless to debate which person likes it more/less). The ramp rate for the Q is on our website, however it is much more indicative of machine performance to compare the time taken to complete a qPCR run
  • How fast can the Q complete a run?
    While this will vary according to your particular assay (different annealing temperatures and hold times), the Q can typically perform a 35 cycle qPCR run complete with melt curve in under 25 min.
  • Why is the efficiency presented as a percentage and not a number between 1→2?
    qPCR efficiency reflects the similarity of data to exponential growth, or a doubling of signal for each qPCR cycle. The efficiency can be expressed as the rate of change of the data (as a multiplication factor), with 1 representing no change in data per cycle and 2 representing a doubling of the data per cycle. Alternatively, this can be expressed as a percentage of amplification efficiency, with 0% representing no change in the data per cycle and 100% representing a doubling of data per cycle.
  • What are the benefits of using the REST analysis rather than ∆∆Ct?
    The REST analysis has the key advantage that it incorporates reaction efficiency while calculating expression ratios (Pfaffl 2002). The inability for the 2-∆∆Ct method to incorporate reaction efficiency could lead to major biases in the calculated expression ratios (eg see Ruijter 2009). While the 2-∆∆Ct method has been modified in some cases to account for amplification efficiencies derived from serial dilution of a standard sample, the combination of linREGPCR and REST allows the reaction efficiency of each actual sample to be used, eliminating the bias of the standard curve method that is often referred to as being only assay specific, and not sample specific (the standard curve method assumes that the calculated efficiency is both a perfect derivation of that value, and that the value perfectly matches that of the actual samples in each instance. The linREGPCR method includes sample specific differences in efficiency and does not assume equivalence of a standard curve with observed values, increasing the accuracy of the calculated data). Finally, the REST analysis uses non-parametric randomisation to calculate the error, which is more suitable for qPCR data than more standard parametric tests (eg T-test). The REST randomisation retains the same power as parametric testing without being affected by data distribution, unlike parametric tests which rely on normal distribution of data which is often not true of qPCR data, and also requires equal variance among the datasets which non-parametric testing does not.
  • Why did you choose linREGPCR for data analysis?
    LinREGPCR uses sophisticated algorithms to increase the accuracy and reproducibility of qPCR data analysis, and is in our opinion the all-round best performing qPCR cycling analysis method. For more information, see (Ramakers 2003, ruijter 2009). The algorithm places the threshold value within the best portion of collected data (the window of linearity), and can scan for the threshold value with the least variation among and between samples to increase the accuracy of the data analysis. Importantly, linREGPCR will calculate efficiency values for each qPCR curve in the data analysis (meaning you don’t need to assume equal efficiency between standards and unknowns, and any sample specific efficiency differences are accounted for). linREGPCR employs a superior baseline correction method that has been shown to reduce the error in efficiency values that can be caused by using other baseline correction methods. Finally, the linREGPCR cycling analysis functions synergistically with our relative quantification analysis to provide the most accurate and reproducible gene expression ratio calculation.
  • If my computer has an issue during a run – how can I retrieve my data?
    If a computer loses communication with the Q, the data from that run is stored on the machine. If you are using the same computer to reconnect to the Q-qPCR, clicking on the serial number icon (top right hand corner of the software) will give an option to ‘recover the run’ file. Similarly, if you connect to the Q-qPCR with a separate computer, you can still recover the data by clicking on the serial number icon, but now the option provided will be to ‘reconstruct the run’.
  • Which fluorophores can I use with the Q?
    The Q-qPCR’s optical detectors will work optimally with FAM (green), Cal Fluor Orange 560 (yellow detector), Cal Fluor Red 610 (orange channel) and Quasar 670 (red channel). This combination of fluorophores will produce less than 3% crosstalk between any/all channel/s used. Other fluorophores with similar spectral properties will also be suitable. For example, Texas Red/ROX have very similar spectral properties to Cal Fluor Red 610, and can be measure on the orange channel. A list of popular fluorophores that are suitable for a particular detector are available while setting up the assay chemistry, and a more comprehensive chart is provided in the Q-qPCR manual (page 143).
  • Can I export the raw data?
    Yes. To export the raw data, expand the save as icon and select the save as excel workbook option. This will save all of the information in the run file, including the raw data.
  • Do I really need water balance tubes?
    Yes, for optimal performance the Q-qPCR should have 48x reactions of equal volume (samples or H20). This is because the amount of material that you need to heat/cool (ie the thermal load) will affect the amount of energy required to heat it to a particular temperature. As an example, a kettle filled with 1000 mL water will take more energy to heat to boiling than that same kettle filled with 100 mL. The Q-qPCR will run to specification when using a fully stacked rotor, but changes to the thermal load (incompletely filling the rotor) may impact the thermal performance.
  • Can the Q-qPCR perform HRM analysis?
    Yes. The Q-qPCR has excellent thermal accuracy (uniformity of ±0.05oC) and can run a melt curve at 0.025oC/sec.
  • Can the samples be used for post-PCR analysis when the tubes have the oil overlay?
    Yes, you can break through the oil layer with a pipette tip to remove your PCR reaction without the oil.
  • What level of ROX should I use?
    The Q-qPCR does not require ROX. Other qPCR machines may require ROX to track temperature and/or optical performance due to thermal drift and variation in the detector setup. The use of magnetic induction (which has no thermal drift and will continue to provide uniform thermal performance over time), combined with the lack of moving parts in the detectors means that the Q-qPCR machine is highly reproducible over time. This means that while other machines may require the ROX channel just to monitor machine performance, the Q-qPCR has the advantage of allowing all channels to be available for real samples.
  • Can I use the Q-qPCR for fieldwork?
    The Q-qPCR is well suited for fieldwork. You can run the machine from any power source, using a power inverter as required. Because you don’t need to calibrate for optics or temperature, and because of its small size/weight, the Q is ideal to transport to field sites while retaining functionality – it stays plug-and-play when other machines would need calibration. Also see our Technical Note: Q in the Field - Alternative Power Sources
  • Can I configure the Q-qPCR software to easily enter sample names from a 96-well plate?
    Yes, the left hand side of the 96-well plate view will list the samples in the A1::H6 format, and the icon beside that will list the samples in the A7::H12 format to match the right hand side of a plate (the A1::H6 and A7::H12 icons are located at the top right hand corner of the samples editor). Next to these icons are the ’samples by column’ and ‘samples by row’ icons. If the samples were originally oriented according to the columns of a 96-well plate then you can simply enter these in to the Q-PCR software as it is the default display. If the samples were originally oriented across the rows of the 96-well plate, the samples will be located in different positions in the rotor, but once you select the ‘samples by row’ icon the samples selector will re-orient the display so that you can easily enter the sample names
  • Can I save my analysis settings to be automatically applied in subsequent runs?
    Yes. You can either click the ‘save analysis settings’ icon which is located at the top right hand corner of the analysis settings display (looks like an Erlenmeyer flask), or expand the assay in the navigation bar (left hand strip of the software) and click the + icon next to the analysis settings icon, fill out your information in the appropriate analysis setting and save (click on the assay name in the navigation pane and click the save icon).
  • How can I find help about the Q-qPCR?
    The best initial resource is the Q-qPCR manual. You can access the manual by clicking on the ‘?’ icon located at the top left hand side of the software, and selecting the Q-qPCR manual option. Once open, you can scroll to the relevant section or use the ctrl+F search function to find what you need.
  • Click here to see all FAQs

    Publications

    Engineered CHO cells as a novel AAV production platform for gene therapy delivery
    Abdou Nagy - 2023
    Abstract
    The Herpes simplex virus (HSV)-based platform for production of recombinant adeno-associated viral vectors (rAAVs) yields higher titers and increased percentage of full capsids when compared to the triple transient transfection (TTT) method. However, this platform currently faces two major challenges. The first challenge is the reliance on commercial media, sometimes supplemented with serum, leading to costly manufacturing and a high risk for introduction of adventitious agents. The second challenge is that the production of HSV-1 relies on adherent complementing Vero cells (V27), making it difficult to scale up. We engineered serum-free-adapted CHO cells expressing key HSV-1 entry receptors, HVEM and/or Nectin-1 to address the first challenge. Using high-throughput cloning methods, we successfully selected a HVEM receptor-expressing clone (CHO–HV–C1) that yields 1.62 × 109, 2.51 × 109, and 4.07 × 109 viral genome copies/mL with rAAV6.2-GFP, rAAV8-GFP, and rAAV9-GFP vectors respectively, within 24 h post rHSV-1 co-infection. Moreover, CHO–HV–C1-derived rAAVs had comparable in vitro transduction, infectivity, and biodistribution titers to those produced by TTT. The second challenge was addressed via engineering CHO–HV–C1 cells to express HSV-1 CP27. These cells successfully produced rHSV-1 vectors, but with significantly lower titers than V27 cells. Taken together, the CHO/HSV system provides a novel, scalable, reduced cost, serum-free AAV manufacturing platform.
    Analytical sensitivity of a method is critical in detection of low-level BRCA1 constitutional epimutation
    Filip Machaj - 2023
    Abstract
    Recent reports based on a substantial number of cases, warrant need for population-based research to determine implications of constitutional methylation of tumor suppressor genes such as BRCA1 occurring in healthy tissue in the prediction of cancer. However, the detection of the constitutional methylation in DNA extracted from blood has already been shown to be technologically challenging, mainly because epimutations appear to be present in blood at a very low level. The analytical sensitivity required for low-level methylation detection can be provided by NGS, but this technique is still labor and cost-intensive. We assessed if PCR-based MS-HRM and BeadChip microarray technologies, which are standardized and cost-effective technologies for methylation changes screening, provide a sufficient level of analytical sensitivity for constitutional BRCA1 methylation detection in blood samples. The study included whole blood samples from 67 healthy women, 35 with previously confirmed and 32 with no detectable BRCA1 promoter methylation for which we performed both MS-HRM based BRCA1 gene methylation screening and genome wide methylation profiling with EPIC microarray. Our results shown, that low-level BRCA1 methylation can be effectively detected in DNA extracted from blood by PCR-based MS-HRM. At the same time, EPIC microarray does not provide conclusive results to unambiguously determine the presence of BRCA1 constitutional methylation in MS-HRM epimutation positive samples. The analytical sensitivity of MS-HRM is sufficient to detect low level BRCA1 constitutional epimutation in DNA extracted from blood and BeadChip technology-based microarrays appear not to provide that level of analytical sensitivity.
    Advances in Malaria Testing: Screening and Identification of Carriers from Saliva
    Sean Campos - 2023
    Abstract
    Plasmodium is a parasite that can infect red blood cells and cause flu-like symptoms with malaria infection. Traditional diagnostic methods do not include counting or testing for gametocytes, which can reservoir in the liver for long periods of time and recirculate. These carriers may have no symptoms, but they can transmit infection to others or to mosquitos. Currently, no diagnostic tests have been approved to detect Plasmodium gametocytes in either symptomatic or asymptomatic whole blood samples. Therefore, we developed real-time PCR assays to detect active and carrier states of malaria. The first is a traditional screening test that can detect any of the five Plasmodium species that cause malaria infection. The second is a companion test to differentiate and quantitate Plasmodium falciparum and P. vivax gametocytes in samples of whole blood from patients who may be asymptomatic and present negative results from screening tests. The screening test showed amplification of P. falciparum, P. vivax, P. ovale, P. malariae, and P. knowlesi in saliva with an overall detection limit of 565 copies/µL. The gametocyte test showed no cross-reactivity between P. falciparum and P. vivax with a limit of detection of RNA at 1000 copies/µL.
    Immunotherapy with IL12 and PD1/CTLA4 inhibition is effective in advanced ovarian cancer and associates with reversal of myeloid cell-induced immunosuppression
    Paul G. Pavicic Jr. - 2023
    Abstract
    The tumor microenvironment (TME) in ovarian cancer (OC) is characterized by immune suppression, due to an abundance of suppressive immune cells populations. To effectively enhance the activity of immune checkpoint inhibition (ICI), there is a need to identify agents that target these immunosuppressive networks while promoting the recruitment of effector T cells into the TME. To this end, we sought to investigate the effect of the immunomodulatory cytokine IL12 alone or in combination with dual-ICI (anti-PD1 + anti-CTLA4) on anti-tumor activity and survival, using the immunocompetent ID8-VEGF murine OC model. Detailed immunophenotyping of peripheral blood, ascites, and tumors revealed that durable treatment responses were associated with reversal of myeloid cell-induced immune suppression, which resulted in enhanced anti-tumor activity by T cells. Single cell transcriptomic analysis further demonstrated striking differences in the phenotype of myeloid cells from mice treated with IL12 in combination with dual-ICI. We also identified marked differences in treated mice that were in remission compared to those whose tumors progressed, further confirming a pivotal role for the modulation of myeloid cell function to allow for response to immunotherapy. These findings provide the scientific basis for the combination of IL12 and ICI to improve clinical response in OC.
    Leptospira enrichment culture followed by ONT metagenomic sequencing allows better detection of Leptospira presence and diversity in water and soil samples
    Myranda Gorman - 2022
    Abstract
    Background Leptospirosis, a life-threatening disease in humans and animals, is one of the most widespread global zoonosis. Contaminated soil and water are the major transmission sources in humans and animals. Clusters of disease outbreaks are common during rainy seasons. Methodology/Principal findings In this study, to detect the presence of Leptospira, we applied PCR, direct metagenomic sequencing, and enrichment culture followed by PCR and metagenomic sequencing on water and soil samples. Direct sequencing and enrichment cultures followed by PCR or sequencing effectively detected pathogenic and nonpathogenic Leptospira compared to direct PCR and 16S amplification-based metagenomic sequencing in soil or water samples. Among multiple culture media evaluated, Ellinghausen-McCullough-Johnson-Harris (EMJH) media containing antimicrobial agents was superior in recovering and detecting Leptospira from the environmental samples. Our results show that enrichment culture followed by PCR can be used to confirm the presence of pathogenic Leptospira in environmental samples. Additionally, metagenomic sequencing on enrichment cultures effectively detects the abundance and diversity of Leptospira spp. from environmental samples. Conclusions/Significance The selection of methodology is critical when testing environmental samples for the presence of Leptospira. Selective enrichment culture improves Leptospira detection efficacy by PCR or metagenomic sequencing and can be used successfully to understand the presence and diversity of pathogenic Leptospira during environmental surveillance.
    Identification of factors associated with Fasciola hepatica infection risk areas on pastures via an environmental DNA survey of Galba truncatula distribution using droplet digital and quantitative real-time PCR assays
    Rhys Aled Jones - 2022
    Abstract
    Environmental DNA (eDNA) is a powerful tool for identifying the spatial and temporal presence and density of species in a range of aquatic habitats. The analysis of eDNA has a wide range of application, one of which may be to inform of Fasciola hepatica infection risk on pastures based on the detection of its eDNA as well as that of its intermediate snail host, Galba truncatula eDNA. Here, droplet digital PCR (ddPCR) and quantitative real-time PCR (qPCR) assays were developed to detect the eDNA of F. hepatica, and its intermediate snail host, G. truncatula in water samples collected from pastures grazed by cattle and/or sheep. Environmental factors associated with species presence, as detected via an eDNA survey, were identified using zero-inflated linear mixed models. Sixty-four habitats were sampled across six farms in Ceredigion, Wales, UK, with ddPCR and qPCR identifying 42 and 33 habitats to be positive for G. truncatula eDNA, respectively. G. truncatula eDNA was significantly less likely to be detected in habitats fully shaded by trees, those that contained black or dark brown soils and habitats that contained deep water pools (p < 0.05). Significantly higher G. truncatula eDNA concentrations were observed in habitats that tend to dry up during Summer (i.e., temporary habitats) (p < 0.05). ddPCR also identified five habitats to be positive for F. hepatica eDNA; however, questions remain regarding the utility of F. hepatica eDNA detection due to a lack of specificity toward infective F. hepatica larval stages. The results of this study inform of factors which influences G. truncatula distribution and ecology on pastures and also provided practical information for farmers to aid F. hepatica control in their flocks and herds.
    A Mobile Laboratory Enables Fecal Pollution Source Tracking in Catchments Using Onsite qPCR Assays
    Rixia Zan - 2022
    Abstract
    Onsite molecular diagnostics can revolutionize fecal pollution source tracking. We aimed to validate a method for onsite qPCR assays with a miniature speaker-sized Q qPCR instrument and other portable equipment items. We showed that marker genes for total bacteria (16S) and E. coli (rodA) in 100 mL of river water measured with this method agreed within ±0.3 log10 units with results obtained when using conventional laboratory equipment items. We then deployed the portable method in a mobile laboratory (‘lab in a van’) and quantified HF183 marker genes for human host associated Bacteroides in river water within 3 h of sampling. We also used the mobile laboratory to investigate urban river water and effluents from two storm drains and a retention pond and collected comprehensive microbial and physicochemical water quality data. We found significantly higher HF183 gene levels in the older storm drain compared to the river water (6.03 ± 0.04 vs. 4.23 ± 0.03 log10 gene copies per 100 mL), and a principal component analysis revealed that storm drain effluent retention in a pond beneficially altered water characteristics, making them more like those of the receiving river. In conclusion, onsite qPCR assays can be performed with portable equipment items to quickly test water.
    Single-tube collection and nucleic acid analysis of clinical samples for SARS-CoV-2 saliva testing
    Kyle H. Cole - 2022
    Abstract
    The SARS-CoV-2 pandemic has brought to light the need for expedient diagnostic testing. Cost and availability of large-scale testing capacity has led to a lag in turnaround time and hindered contact tracing efforts, resulting in a further spread of SARS-CoV-2. To increase the speed and frequency of testing, we developed a cost-effective single-tube approach for collection, denaturation, and analysis of clinical samples. The approach utilizes 1 µL microbiological inoculation loops to collect saliva, sodium dodecyl sulfate (SDS) to inactivate and release viral genomic RNA, and a diagnostic reaction mix containing polysorbate 80 (Tween 80). In the same tube, the SDS-denatured clinical samples are introduced to the mixtures containing all components for nucleic acids detection and Tween 80 micelles to absorb the SDS and allow enzymatic reactions to proceed, obviating the need for further handling of the samples. The samples can be collected by the tested individuals, further decreasing the need for trained personnel to administer the test. We validated this single-tube sample-to-assay method with reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) and reverse transcription loop-mediated isothermal amplification (RT-LAMP) and discovered little-to-no difference between Tween- and SDS-containing reaction mixtures, compared to control reactions. This approach reduces the logistical burden of traditional large-scale testing and provides a method of deployable point-of-care diagnostics to increase testing frequency.
    Single-tube collection and nucleic acid analysis of clinical samples for SARS-CoV-2 saliva testing
    Kyle H. Cole - 2022
    Abstract
    The SARS-CoV-2 pandemic has brought to light the need for expedient diagnostic testing. Cost and availability of large-scale testing capacity has led to a lag in turnaround time and hindered contact tracing efforts, resulting in a further spread of SARS-CoV-2. To increase the speed and frequency of testing, we developed a cost-effective single-tube approach for collection, denaturation, and analysis of clinical samples. The approach utilizes 1 µL microbiological inoculation loops to collect saliva, sodium dodecyl sulfate (SDS) to inactivate and release viral genomic RNA, and a diagnostic reaction mix containing polysorbate 80 (Tween 80). In the same tube, the SDS-denatured clinical samples are introduced to the mixtures containing all components for nucleic acids detection and Tween 80 micelles to absorb the SDS and allow enzymatic reactions to proceed, obviating the need for further handling of the samples. The samples can be collected by the tested individuals, further decreasing the need for trained personnel to administer the test. We validated this single-tube sample-to-assay method with reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) and reverse transcription loop-mediated isothermal amplification (RT-LAMP) and discovered little-to-no difference between Tween- and SDS-containing reaction mixtures, compared to control reactions. This approach reduces the logistical burden of traditional large-scale testing and provides a method of deployable point-of-care diagnostics to increase testing frequency.
    Dissolved Microcystin Release Coincident with Lysis of a Bloom Dominated by Microcystis spp. in Western Lake Erie Attributed to a Novel Cyanophage
    Katelyn M. McKindles - 2020
    Abstract
    Western Lake Erie (Laurentian Great Lakes) is prone to annual cyanobacterial harmful algal blooms (cHABs) dominated by Microcystis spp. that often yield microcystin toxin concentrations exceeding the federal EPA recreational contact advisory of 8 μg liter−1. In August 2014, microcystin levels were detected in finished drinking water above the World Health Organization 1.0 μg liter−1 threshold for consumption, leading to a 2-day disruption in the supply of drinking water for >400,000 residents of Toledo, Ohio (USA). Subsequent metatranscriptomic analysis of the 2014 bloom event provided evidence that release of toxin into the water supply was likely caused by cyanophage lysis that transformed a portion of the intracellular microcystin pool into the dissolved fraction, rendering it more difficult to eliminate during treatment. In August 2019, a similar increase in dissolved microcystins at the Toledo water intake was coincident with a viral lytic event caused by a phage consortium different in composition from what was detected following the 2014 Toledo water crisis. The most abundant viral sequence in metagenomic data sets was a scaffold from a putative member of the Siphoviridae, distinct from the Ma-LMM01-like Myoviridae that are typically documented to occur in western Lake Erie. This study provides further evidence that viral activity in western Lake Erie plays a significant role in transformation of microcystins from the particulate to the dissolved fraction and therefore requires monitoring efforts from local water treatment plants. Additionally, identification of multiple lytic cyanophages will enable the development of a quantitative PCR toolbox to assess viral activity during cHABs. IMPORTANCE Viral attack on cHABs may contribute to changes in community composition during blooms, as well as bloom decline, yet loss of bloom biomass does not eliminate the threat of cHAB toxicity. Rather, it may increase risks to the public by delivering a pool of dissolved toxin directly into water treatment utilities when the dominating Microcystis spp. are capable of producing microcystins. Detecting, characterizing, and quantifying the major cyanophages involved in lytic events will assist water treatment plant operators in making rapid decisions regarding the pool of microcystins entering the plant and the corresponding best practices to neutralize the toxin.
    Controlled Release in Hydrogels Using DNA Nanotechnology
    Chih-Hsiang Hu - 2022
    Abstract
    Gelatin is a biopolymer widely used to synthesize hydrogels for biomedical applications, such as tissue engineering and bioinks for 3D bioprinting. However, as with other biopolymer-based hydrogels, gelatin-hydrogels do not allow precise temporal control of the biomolecule distribution to mimic biological signals involved in biological mechanisms. Leveraging DNA nanotechnology tools to develop a responsive controlled release system via strand displacement has demonstrated the ability to encode logic process, which would enable a more sophisticated design for controlled release. However, this unique and dynamic system has not yet been incorporated within any hydrogels to create a complete release circuit mechanism that closely resembles the sequential distribution of biomolecules observed in the native environment. Here, we designed and synthesized versatile multi-arm DNA motifs that can be easily conjugated within a gelatin hydrogel via click chemistry to incorporate a strand displacement circuit. After validating the incorporation and showing the increased stability of DNA motifs against degradation once embedded in the hydrogel, we demonstrated the ability of our system to release multiple model cargos with temporal specificity by the addition of the trigger strands specific to each cargo. Additionally, we were able to modulate the rate and quantity of cargo release by tuning the sequence of the trigger strands.
    Activation of LXR Receptors and Inhibition of TRAP1 Causes Synthetic Lethality in Solid Tumors
    Trang Thi Thu Nguyen - 2019
    Abstract
    Cholesterol is a pivotal factor for cancer cells to entertain their relentless growth. In this case, we provide a novel strategy to inhibit tumor growth by simultaneous activation of liver-X-receptors and interference with Tumor Necrosis Factor Receptor-associated Protein 1 (TRAP1). Informed by a transcriptomic and subsequent gene set enrichment analysis, we demonstrate that inhibition of TRAP1 results in suppression of the cholesterol synthesis pathway in stem-like and established glioblastoma (GBM) cells by destabilizing the transcription factor SREBP2. Notably, TRAP1 inhibition induced cell death, which was rescued by cholesterol and mevalonate. Activation of liver X receptor (LXR) by a clinically validated LXR agonist, LXR623, along with the TRAP1 inhibitor, gamitrinib (GTPP), results in synergistic reduction of tumor growth and cell death induction in a broad range of solid tumors, which is rescued by exogenous cholesterol. The LXR agonist and TRAP1 inhibitor mediated cell death is regulated at the level of Bcl-2 family proteins with an elevation of pro-apoptotic Noxa. Silencing of Noxa and its effector BAK attenuates cell death mediated by the combination treatment of LXR agonists and TRAP1 inhibition. Combined inhibition of TRAP1 and LXR agonists elicits a synergistic activation of the integrated stress response with an increase in activating transcription factor 4 (ATF4) driven by protein kinase RNA-like endoplasmic reticulum kinase (PERK). Silencing of ATF4 attenuates the increase of Noxa by using the combination treatment. Lastly, we demonstrate in patient-derived xenografts that the combination treatment of LXR623 and gamitrinib reduces tumor growth more potent than each compound. Taken together, these results suggest that TRAP1 inhibition and simultaneous activation of LXR might be a potent novel treatment strategy for solid malignancies.
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