STAMP: a multiplex sequencing method for simultaneous evaluation of mitochondrial DNA heteroplasmies and content

ABSTRACTHuman mitochondrial genome (mtDNA) variations, such as mtDNA heteroplasmies (the co-existence of mutated and wild-type mtDNA), have received increasing attention in recent years for their clinical relevance to numerous diseases. But large-scale population studies of mtDNA heteroplasmies have been lagging due to the lack of a labor- and cost-effective method. Here, we present a novel human mtDNA sequencing method called STAMP (sequencing by targeted amplification of multiplex probes) for measuring mtDNA heteroplasmies and content in a streamlined workflow. We show that STAMP has high mapping rates to mtDNA, deep coverage of unique reads, and high tolerance to sequencing and PCR errors when applied to human samples. STAMP also has high sensitivity and low false positive rates in identifying artificial mtDNA variants at fractions as low as 0.5% in genomic DNA samples. We further extend STAMP, by including nuclear DNA-targeting probes, to enable assessment of relative mtDNA content in the same assay. The high cost-effectiveness of STAMP, along with the flexibility of using it for measuring various aspects of mtDNA variations, will accelerate the research of mtDNA heteroplasmies and content in large population cohorts, and in the context of human diseases and aging.

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STAMP: a multiplex sequencing method for simultaneous evaluation of mitochondrial DNA heteroplasmies and content

NAR Genomics and Bioinformatics ◽

10.1093/nargab/lqaa065 ◽

2020 ◽

Vol 2 (4) ◽

Author(s):

Xiaoxian Guo ◽

Yiqin Wang ◽

Ruoyu Zhang ◽

Zhenglong Gu

Keyword(s):

Large Scale ◽

Nuclear Dna ◽

Large Population ◽

High Sensitivity ◽

Cost Effective ◽

Cost Effective Method ◽

Sequencing Method ◽

Scale Population ◽

Mtdna Variants ◽

Multiplex Sequencing

Abstract Human mitochondrial genome (mtDNA) variations, such as mtDNA heteroplasmies (the co-existence of mutated and wild-type mtDNA), have received increasing attention in recent years for their clinical relevance to numerous diseases. But large-scale population studies of mtDNA heteroplasmies have been lagging due to the lack of a labor- and cost-effective method. Here, we present a novel human mtDNA sequencing method called STAMP (sequencing by targeted amplification of multiplex probes) for measuring mtDNA heteroplasmies and content in a streamlined workflow. We show that STAMP has high-mapping rates to mtDNA, deep coverage of unique reads and high tolerance to sequencing and polymerase chain reaction errors when applied to human samples. STAMP also has high sensitivity and low false positive rates in identifying artificial mtDNA variants at fractions as low as 0.5% in genomic DNA samples. We further extend STAMP, by including nuclear DNA-targeting probes, to enable assessment of relative mtDNA content in the same assay. The high cost-effectiveness of STAMP, along with the flexibility of using it for measuring various aspects of mtDNA variations, will accelerate the research of mtDNA heteroplasmies and content in large population cohorts, and in the context of human diseases and aging.

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Fabrication and Characterization of Humidity Sensors Based on Graphene Oxide–PEDOT:PSS Composites on a Flexible Substrate

Micromachines ◽

10.3390/mi11020148 ◽

2020 ◽

Vol 11 (2) ◽

pp. 148 ◽

Cited By ~ 6

Author(s):

Francisco J. Romero ◽

Almudena Rivadeneyra ◽

Markus Becherer ◽

Diego P. Morales ◽

Noel Rodríguez

Keyword(s):

Graphene Oxide ◽

Large Scale ◽

Flexible Substrate ◽

High Sensitivity ◽

Cost Effective ◽

Humidity Sensors ◽

Good Thermal Stability ◽

Cost Effective Method ◽

And Performance ◽

Stability Time

In this paper, we present a simple, fast, and cost-effective method for the large-scale fabrication of high-sensitivity humidity sensors on flexible substrates. These sensors consist of a micro screen-printed capacitive structure upon which a sensitive layer is deposited. We studied two different structures and three different sensing materials by modifying the concentration of poly(3,4-ethylenedioxythiophene)/polystyrene sulfonate (PEDOT:PSS) in a graphene oxide (GO) solution. The results show that the aggregation of the PEDOT:PSS to the GO can modify its electrical properties, boosting the performance of the capacitive sensors in terms of both resistive losses and sensitivity to relative humidity (RH) changes. Thus, in an area less than 30 mm2, the GO/PEDOT:PSS-based sensors can achieve a sensitivity much higher (1.22 nF/%RH at 1 kHz) than other similar sensors presented in the literature which, together with their good thermal stability, time response, and performance over bending, demonstrates that the manufacturing approach described in this work paves the way for the mass production of flexible humidity sensors in an inexpensive way.

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A Short-Read Multiplex Sequencing Method for Reliable, Cost-Effective and High-Throughput Genotyping in Large-Scale Studies

Human Mutation ◽

10.1002/humu.22486 ◽

2013 ◽

Vol 35 (2) ◽

pp. 270-270 ◽

Cited By ~ 1

Author(s):

Hongzhi Cao ◽

Yu Wang ◽

Wei Zhang ◽

Xianghua Chai ◽

Xiandong Zhang ◽

...

Keyword(s):

High Throughput ◽

Large Scale ◽

Cost Effective ◽

Short Read ◽

Sequencing Method ◽

Multiplex Sequencing

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A Short-Read Multiplex Sequencing Method for Reliable, Cost-Effective and High-Throughput Genotyping in Large-Scale Studies

Human Mutation ◽

10.1002/humu.22439 ◽

2013 ◽

Vol 34 (12) ◽

pp. 1715-1720 ◽

Cited By ~ 34

Author(s):

Hongzhi Cao ◽

Yu Wang ◽

Wei Zhang ◽

Xianghua Chai ◽

Xiandong Zhang ◽

...

Keyword(s):

High Throughput ◽

Large Scale ◽

Cost Effective ◽

Short Read ◽

Sequencing Method ◽

Multiplex Sequencing

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Improving COVID-19 Testing Efficiency using Guided Agglomerative Sampling

10.1101/2020.04.13.039792 ◽

2020 ◽

Author(s):

Fayyaz Minhas ◽

Dimitris Grammatopoulos ◽

Lawrence Young ◽

Imran Amin ◽

David Snead ◽

...

Keyword(s):

Large Scale ◽

Resource Constraints ◽

Large Population ◽

Sampling Strategy ◽

Population Surveillance ◽

Large Scale Testing ◽

Scale Population ◽

Simulation Results ◽

The Given ◽

Test Outcomes

AbstractOne of the challenges in the current COVID-19 crisis is the time and cost of performing tests especially for large-scale population surveillance. Since, the probability of testing positive in large population studies is expected to be small (<15%), therefore, most of the test outcomes will be negative. Here, we propose the use of agglomerative sampling which can prune out multiple negative cases in a single test by intelligently combining samples from different individuals. The proposed scheme builds on the assumption that samples from the population may not be independent of each other. Our simulation results show that the proposed sampling strategy can significantly increase testing capacity under resource constraints: on average, a saving of ~40% tests can be expected assuming a positive test probability of 10% across the given samples. The proposed scheme can also be used in conjunction with heuristic or Machine Learning guided clustering for improving the efficiency of large-scale testing further. The code for generating the simulation results for this work is available here: https://github.com/foxtrotmike/AS.

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Coupling multiplex pre-amplification and droplet digital PCR for longitudinal monitoring ofESR1andPIK3CAmutations from plasma cell-free DNA

10.1101/598847 ◽

2019 ◽

Author(s):

Huilan Yao ◽

Grant Wu ◽

Subhasree Das ◽

Crystal MacKenzie ◽

Hua Gao ◽

...

Keyword(s):

Limit Of Detection ◽

Hot Spot ◽

Metastatic Breast ◽

High Sensitivity ◽

Cost Effective ◽

Digital Pcr ◽

Cost Effective Method ◽

High Prevalence ◽

High Concordance ◽

Hotspot Mutations

AbstractHere we report on the development of a sensitive and cost-effective method to longitudinally trackESR1andPIK3CAmutations from cfDNA in patients with metastatic breast cancer (MBC) using a streamlined and de-centralized workflow. Hotspot mutations inESR1have been shown to cause resistance to aromatase inhibitor–based and anti-estrogenic therapies, whilePIK3CAmutations have high prevalence in MBC. As a result, their utility as circulating biomarkers to predict or monitor response in the clinical development of investigational compounds has been the focus of many studies. Six regions inESR1andPIK3CAgenes containing 20 hotspot mutations were pre-amplified, followed by optimized singleplex ddPCR assays to detect allele frequencies of individual mutations. Without pre-amplification, the limit of detection (LOD) and limit of linearity (LOL) of individual ddPCR assays were at 0.05-0.1% and 0.25% level, respectively. With pre-amplification, the LOD and LOL were slightly elevated at 0.1-0.25% and 0.25-0.5% levels, respectively. High concordance was achieved to the BEAMing assay (Sysmex Inostics) for mutation positive assays (r=0.98, P<0.0001). In conclusion, coupling pre-amplification and ddPCR assays allowed us for the detection of up to 20 hot spot mutations inESR1andPIK3CAwith high sensitivity and reproducibility.

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Cost-effectiveness of two monitoring strategies for the great crested newt (Triturus cristatus)

Amphibia-Reptilia ◽

10.1163/156853810791769374 ◽

2010 ◽

Vol 31 (3) ◽

pp. 403-410 ◽

Cited By ~ 3

Author(s):

Jérôme Pellet ◽

Madeleine Kröpfli ◽

Patrick Heer

Keyword(s):

Large Scale ◽

Cost Effective ◽

Amphibian Conservation ◽

Submerged Vegetation ◽

Vegetation Density ◽

Triturus Cristatus ◽

Monitoring Strategies ◽

Cost Effective Method ◽

Crested Newt ◽

Monitoring Protocols

AbstractDesigning cost-effective monitoring protocols is a fundamental prerequisite for amphibian conservation. Here, we report a comparison of flashlight survey and trapping (with and without light sticks as trap baits) in order to determine flashlight detectability and trap detectability of great crested newts (Triturus cristatus). Twelve ponds were surveyed in Switzerland where T. cristatus had been known to occur. We measured covariates affecting both flashlight detectability and trap detectability. Newt flashlight detectability using 20 min long flashlight surveys was on average ± SE = 39% ± 10%). Flashlight detectability was mostly influenced by surface and submerged vegetation density, as well as by water temperature. Newt trap detectability during one night using six funnel traps per pond was on average±SE = 41%±10%. Trap detectability was mainly affected by trap position in the pond, with traps lying on the pond floor being more likely to attract newts. The use of light sticks did not enhance the trap detectability. Estimates of flashlight detectability and trap detectability were used to define how many times the sites have to be visited to be 95% certain of not missing T. cristatus in ponds where they are present. In both cases multiple visits (7 flashlight surveys or 6 trapping sessions) have to be performed. Flashlight surveys are the most easily applied and most cost-effective method to use in large scale programs.

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High efficient extracellular production of recombinant Streptomyces PMF phospholipase D in Escherichia coli

10.21203/rs.3.rs-18976/v1 ◽

2020 ◽

Author(s):

Jing Wang ◽

Sheng Xu ◽

Yang Pang ◽

Xin Wang ◽

Kequan Chen ◽

...

Keyword(s):

Phospholipase D ◽

Signal Peptide ◽

Large Scale ◽

Expression System ◽

Cost Effective ◽

Scale Production ◽

Secretory Expression ◽

E Coli ◽

Cost Effective Method ◽

High Efficient

Abstract Background Currently, Streptomyces is widely used in the preparation of phospholipase D (PLD) with high transphosphatidylation activity. However, the yield of PLD from Streptomyces was low and the culture period was long. Therefore, an efficient and cost-effective method is needed urgently.Results Firstly, PLDs from Streptomyces PMF and Streptomyces racemochromogenes were separately over-expressed in E. coli to compare their transphosphatidylation activity based on the synthesis of phosphatidylserine (PS), and PLDPMF was determined to have higher activity. To further improve PLDPMF synthesis, a secretory expression system suitable for PLDPMF was constructed and optimized with diﬀerent signal peptides. The highest secretory efficiency was observed when the PLDPMF gene was expressed together with its native signal peptide (Nat) and the signal peptide PelB from E. coli. For the application of recombinant PLD to PS synthesis, the PLD properties were characterized and 30.2 g/L of PS was produced after 24 h of bioconversion when 50 g/L phosphatidylcholine (PC) was added.Conclusions We succeeded in over-expressing PLD from Streptomyces PMF in E. coli with high transphosphatidylation activity and enhanced the yield by secretory expression. The secreted PLD was successfully used in the production of PS. Our work makes the large-scale production of PLD and PS feasible.

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An Overview of the Photocatalytic Water Splitting over Suspended Particles

Catalysts ◽

10.3390/catal11010060 ◽

2021 ◽

Vol 11 (1) ◽

pp. 60

Author(s):

Muhammad Amtiaz Nadeem ◽

Mohd Adnan Khan ◽

Ahmed Abdeslam Ziani ◽

Hicham Idriss

Keyword(s):

Water Splitting ◽

Large Scale ◽

Cost Effective ◽

Synthetic Methods ◽

Scale Production ◽

Practical Applications ◽

Co Catalysts ◽

Large Scale Production ◽

Cost Effective Method ◽

Order Of Magnitude

The conversion of solar to chemical energy is one of the central processes considered in the emerging renewable energy economy. Hydrogen production from water splitting over particulate semiconductor catalysts has often been proposed as a simple and a cost-effective method for large-scale production. In this review, we summarize the basic concepts of the overall water splitting (in the absence of sacrificial agents) using particulate photocatalysts, with a focus on their synthetic methods and the role of the so-called “co-catalysts”. Then, a focus is then given on improving light absorption in which the Z-scheme concept and the overall system efficiency are discussed. A section on reactor design and cost of the overall technology is given, where the possibility of the different technologies to be deployed at a commercial scale and the considerable challenges ahead are discussed. To date, the highest reported efficiency of any of these systems is at least one order of magnitude lower than that deserving consideration for practical applications.

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