Enrichment Technology of Low-Abundance Proteomics Based on Magnetic Micro-/Nanomaterial

PLoS ONE ◽  
2020 ◽  
Vol 15 (12) ◽  
pp. e0243781
Author(s):  
Jennifer L. Steele ◽  
Richard C. Stevens ◽  
Oscar A. Cabrera ◽  
Gary J. Bassill ◽  
Sabrina M. Cramer ◽  
...  

The programmable sequence specificity of CRISPR has found uses in gene editing and diagnostics. This manuscript describes an additional application of CRISPR through a family of novel DNA enrichment technologies. CAMP (CRISPR Associated Multiplexed PCR) and cCAMP (chimeric CRISPR Associated Multiplexed PCR) utilize the sequence specificity of the Cas9/sgRNA complex to target loci for the ligation of a universal adapter that is used for subsequent amplification. cTRACE (chimeric Targeting Rare Alleles with CRISPR-based Enrichment) also applies this method to use Cas9/sgRNA to target loci for the addition of universal adapters, however it has an additional selection for specific mutations through the use of an allele-specific primer. These three methods can produce multiplex PCR that significantly reduces the optimization required for every target. The methods are also not specific to any downstream analytical platform. We additionally will present a mutation specific enrichment technology that is non-amplification based and leaves the DNA in its native state: TRACE (Targeting Rare Alleles with CRISPR-based Enrichment). TRACE utilizes the Cas9/sgRNA complex to sterically protect the ends of targeted sequences from exonuclease activity which digests both the normal variant as well as any off-target sequences.


2019 ◽  
Vol 10 ◽  
Author(s):  
Lorenzo Barchi ◽  
Alberto Acquadro ◽  
David Alonso ◽  
Giuseppe Aprea ◽  
Laura Bassolino ◽  
...  

2019 ◽  
Vol 24 (5) ◽  
pp. 477-488 ◽  
Author(s):  
Pan Zhang ◽  
Joo Chuan Yeo ◽  
Chwee Teck Lim

Extracellular vesicles (EVs) are lipid bilayer-bound vesicles secreted by cells. Subtypes of EVs such as microvesicles and exosomes are further categorized mainly by their different biogenesis mechanisms. EVs have been revealed to play an important role in disease diagnosis and intercellular communication. Despite the wide interest in EVs, the technologies for the purification and enrichment of EVs are still in their infancy. The isolation of EVs, especially exosomes, is inherently challenging due to their small size and heterogeneity. In this review, we mainly introduce the advances of techniques in isolating microvesicles and exosomes according to their approaches. Also, we discuss the limitations of currently reported technologies in terms of their specificity and efficiency, and provide our thoughts about future developments of EV purification and enrichment technology.


2017 ◽  
Vol 65 (3) ◽  
pp. 327-339 ◽  
Author(s):  
Wenlong Zhang ◽  
Xiaodan Liu ◽  
Mengcheng Liu ◽  
Bo Ma ◽  
Li Xu ◽  
...  

Pasteurella multocida, Mannheimia haemolytica and Trueperella pyogenes are three bacterial pathogens closely associated with the bovine respiratory disease complex (BRDC). In the current study, a multiplex PCR for the simultaneous detection of these three bacteria in cultures was established. After serial optimisation, the detection limit of the method for the genomic DNA of the three bacteria was 40 pg/μl. The method could detect the genomic DNA of these three bacteria but not the genomic DNA of seven other bacterial strains. Together with the bacterial enrichment technology, the multiplex PCR could be used for detecting the three bacteria in animal tissues. This method might be valuable for speeding up laboratory diagnosis and directing the treatment of BRDC to these three bacterial pathogens.


2011 ◽  
Vol 29 (15_suppl) ◽  
pp. 5511-5511 ◽  
Author(s):  
K. Jatana ◽  
P. Balasubramanian ◽  
J. C. Lang ◽  
T. N. Teknos ◽  
D. E. Schuller ◽  
...  

1966 ◽  
Vol 23 (8) ◽  
pp. 437-440
Author(s):  
V. S. Semenov ◽  
V. A. Lipchinskaya

2019 ◽  
Vol 28 (3) ◽  
pp. 537-545
Author(s):  
G. A. Shevchenko ◽  
V. G. Shevchenko ◽  
V. A. Baranov ◽  
V. N. Spassky

The purpose of this research is the development of a technology for the enrichment of slag from metallurgical processing of copper ore concentrate based on     the results of spectral, chemical, sieve and petrographic analysis. The results of spectral analysis indicate the copper content in all three samples of mineral raw materials at more than 1 %. The results of chemical analysis indicated a high copper content in the samples from 13.4 to 17.1%, as well as a high iron content from 9 to 18%. Analysis of the results of the sieve analysis showed that the largest amount of copper is contained in the size classes 0.063–0.05 mm at from 18.6 to 24.1 % and 0.04 mm at from 15.6 to 38 %. In accordance with the petrographic studies, the size of copper grains varies from 0.1–0.3 to 1–5 mm. The most common sizes of copper grains in the studied samples are 0.2-0.3 and 1-2 mm. Based on the results of spectral, chemical, sieve and petrographic analysis, a technology for the enrichment of copper-containing slags has been developed. Gravity wet enrichment technology with a capacity of 5 t/h with Cu recovery in the range of 80–95 % suggests the grinding of raw materials with a constant water supply up to 40 m3/h from the sludge collector. The heavy fractions are fed to a magnetic separator and then to a classifier for the extraction of magnetic concentrate and slag, which after the separation of the fraction of 0.08-0.4 mm with the MVG screen can later be used as a raw material for the building industry. The light fractions after the concentration tables are fed to the classifier, on which the copper concentrate is released. The average density fractions are returned to the closed cycle for further grinding in a ball mill. However, such a wet enrichment scheme requires a continuous water supply and the sludge collector’s presence, which cannot always be ensured. Therefore, the technology of slag dry enrichment with a particle size of 0–100 mm has also been developed. The central apparatus in the proposed enrichment technologies is the MVG vibrating screen, which is designed to separate bulk materials by particle size from 20 microns to several millimeters. Polyfrequency oscillations in the frequency range from several Hz to kHz are implemented on the screen, eliminating blockage of the sieve cells, destruction of the formed aggregates of stuck particles, ensuring their intensive movement in the layer and efficient passage of particles reaching the sieve surface through the cells. This type of vibration makes it possible to achieve much greater efficiency of separation and dehydration of materials than in traditional screens and to ensure continuous self-cleaning of the mesh, which contributes to the process of separation and dehydration. Due to the lack of tension, high durability of the working surface is ensured. Due to the transfer of minimum loads on the base, the screen is installed without arranging special foundations, including on the floors of buildings and structures. A standard- sized row of screens was developed with a screening surface area from 1 to 4 m2 and a different number of tiers.


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