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1.
Anal Chem ; 94(6): 2926-2933, 2022 02 15.
Article in English | MEDLINE | ID: covidwho-1721378

ABSTRACT

Recombinase polymerase amplification (RPA) is a useful pathogen identification method. Several label-free detection methods for RPA amplicons have been developed in recent years. However, these methods still lack sensitivity, specificity, efficiency, or simplicity. In this study, we propose a rapid, highly sensitive, and label-free pathogen assay system based on a solid-phase self-interference RPA chip (SiSA-chip) and hyperspectral interferometry. The SiSA-chips amplify and capture RPA amplicons on the chips, rather than irrelevant amplicons such as primer dimers, and the SiSA-chips are then analysed by hyperspectral interferometry. Optical length increases of SiSA-chips are used to demonstrate RPA detection results, with a limit of detection of 1.90 nm. This assay system can detect as few as six copies of the target 18S rRNA gene of Plasmodium falciparum within 20 min, with a good linear relationship between the detection results and the concentration of target genes (R2 = 0.9903). Single nucleotide polymorphism (SNP) genotyping of the dhfr gene of Plasmodium falciparum is also possible using the SiSA-chip, with as little as 1% of mutant gene distinguished from wild-type loci (m/wt). This system offers a high-efficiency (20 min), high-sensitivity (6 copies/reaction), high-specificity (1% m/wt), and low-cost (∼1/50 of fluorescence assays for RPA) diagnosis method for pathogen DNA identification. Therefore, this system is promising for fast identification of pathogens to help diagnose infectious diseases, including SNP genotyping.


Subject(s)
Nucleic Acid Amplification Techniques , Recombinases , Interferometry , Nucleic Acid Amplification Techniques/methods , Nucleotidyltransferases , Plasmodium falciparum/genetics , Sensitivity and Specificity
2.
Micromachines (Basel) ; 12(12)2021 Dec 19.
Article in English | MEDLINE | ID: covidwho-1580576

ABSTRACT

A two-stage isothermal amplification method, which consists of a first-stage basic recombinase polymerase amplification (RPA) and a second-stage fluorescence loop-mediated isothermal amplification (LAMP), as well as a microfluidic-chip-based portable system, were developed in this study; these enabled parallel detection of multiplex targets in real time in around one hour, with high sensitivity and specificity, without cross-contamination. The consumption of the sample and the reagent was 2.1 µL and 10.6 µL per reaction for RPA and LAMP, respectively. The lowest detection limit (LOD) was about 10 copies. The clinical amplification of about 40 nasopharyngeal swab samples, containing 17 SARS-CoV-2 (severe acute respiratory syndrome coronavirus) and 23 measles viruses (MV), were parallel tested by using the microfluidic chip. Both clinical specificity and sensitivity were 100% for MV, and the clinical specificity and sensitivity were 94.12% and 95.83% for SARS-CoV-2, respectively. This two-stage isothermal amplification method based on the microfluidic chip format offers a convenient, clinically parallel molecular diagnostic method, which can identify different nucleic acid samples simultaneously and in a timely manner, and with a low cost of the reaction reagent. It is especially suitable for resource-limited areas and point-of-care testing (POCT).

3.
Agronomy ; 11(11):2346, 2021.
Article in English | Academic Search Complete | ID: covidwho-1542394

ABSTRACT

The photosynthetically active radiation (PAR) of crop canopy is highly related to yield formation, but how it relates to yield and yield distribution is not well understood. The focus of this study was to explore the relationship between light competition under different densities and yield distributions of cotton. The experiment was conducted in 2019 and 2020 at the Cotton Research Institute of the Chinese Academy of Agricultural Sciences in Anyang city, Henan Province, China. A randomized block design was employed, with a total of three repeats. Each repeat had six density treatments: D1: 15,000;D2: 33,000;D3: 51,000;D4: 69,000;D5: 87,000;and D6: 105,000 plants·ha−1. As predicted, the results showed that the canopy light interception, leaf area index, plant height, and biomass of high-density cotton were higher than those of low-density cotton. The aboveground biomass produced by D6 was the highest, and was 12.9, 19.5, 25.4, 46.3, and 69.2% higher in 2019 and 14.3, 19.9, 32.5, 53.7, and 109.9% higher in 2020 than D5, D4, D3, D2, and D1, respectively. Leaf area, plant height, biomass, boll number, and boll weight were significantly correlated with the light interception rate. D5 (87,000 plants·ha−1) had a higher light interception rate and the highest yield. The highest lint yields produced by D5 were 1673.5 and 1375.4 kg·ha−1 in two years, and was 3.2, 4.3, 5.6, 9.7, and 24.7% higher in 2019, and 6.8, 10.6, 13.5, 21.5, and 34.4% higher in 2020 than D6, D4, D3, D2, and D1, respectively. The boll retention of the lower fruit branch under D5 reached 0.51 and 0.57 in two years, respectively. The shedding rate of the upper fruit branch decreased with the increase in cotton density in two years. The boll retention rate and shedding rate in the lower part of cotton plants were most closely related to light interception, with R2 values of 0.91 and 0.96, respectively. Our study shows cotton yield could be improved through higher light interception by optimizing planting density and canopy structure. [ FROM AUTHOR] Copyright of Agronomy is the property of MDPI and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full . (Copyright applies to all s.)

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