scholarly journals DNA Sequencing Using Rolling Circle Amplification and Precision Glass Syringes in a High-Throughput Liquid Handling System

BioTechniques ◽  
2003 ◽  
Vol 34 (1) ◽  
pp. 204-207 ◽  
Author(s):  
Hui-Chung Wu ◽  
Jean Shieh ◽  
David J. Wright ◽  
Arezou Azarani
Keyword(s):  
Dna Sequencing ◽  
High Throughput ◽  
Rolling Circle Amplification ◽  
Rolling Circle ◽  
Handling System ◽  
Liquid Handling

scholarly journals High-throughput Quant-iT PicoGreen assay using an automated liquid handling system

BioTechniques ◽  
2019 ◽  
Vol 66 (6) ◽  
pp. 290-294 ◽  
Author(s):  
Kay Anantanawat ◽  
Nicola Pitsch ◽  
Caroline Fromont ◽  
Caroline Janitz
Keyword(s):  
High Throughput ◽  
Handling System ◽  
Liquid Handling ◽  
Picogreen Assay

scholarly journals High-throughput, cost-effective verification of structural DNA assembly

2013 ◽  
Vol 42 (4) ◽  
pp. e22-e22 ◽  
Author(s):  
Yandi Dharmadi ◽  
Kedar Patel ◽  
Elaine Shapland ◽  
Daniel Hollis ◽  
Todd Slaby ◽  
...  
Keyword(s):  
High Throughput ◽  
Low Cost ◽  
Rolling Circle Amplification ◽  
Cost Effective ◽  
Building Blocks ◽  
Pathway Engineering ◽  
Dna Assembly ◽  
Rolling Circle ◽  
Enzyme Screening ◽  
First Time

Abstract DNA ‘assembly’ from ‘building blocks’ remains a cornerstone in synthetic biology, whether it be for gene synthesis (∼1 kb), pathway engineering (∼10 kb) or synthetic genomes (>100 kb). Despite numerous advances in the techniques used for DNA assembly, verification of the assembly is still a necessity, which becomes cost-prohibitive and a logistical challenge with increasing scale. Here we describe for the first time a comprehensive, high-throughput solution for structural DNA assembly verification by restriction digest using exhaustive in silico enzyme screening, rolling circle amplification of plasmid DNA, capillary electrophoresis and automated digest pattern recognition. This low-cost and robust methodology has been successfully used to screen over 31 000 clones of DNA constructs at <$1 per sample.

scholarly journals Fully Automated Peptide Mapping Protocol for Multi-Attribute Method by Liquid Chromatography–Tandem Mass Spectroscopy with a High-Throughput Robotic Liquid Handling System

2020 ◽  
Author(s):  
Chen Qian ◽  
Ben Niu ◽  
Rod Brian Jimenez ◽  
Jihong Wang ◽  
Methal Albarghouthi ◽  
...  
Keyword(s):  
Liquid Chromatography ◽  
Mass Spectroscopy ◽  
High Throughput ◽  
Peptide Mapping ◽  
Tandem Mass ◽  
Handling System ◽  
Liquid Handling ◽  
Hands On ◽  
Liquid Chromatography Tandem Mass ◽  
Tandem Mass Spectroscopy

ABSTRACTThe multi-attribute method (MAM) based on liquid chromatography–tandem mass spectroscopy is emerging as a powerful tool to directly monitor multiple product quality attributes simultaneously. Preparation of samples for MAM, however, is labor intensive, involving protein denaturation, disulfide bond reduction, free cysteine alkylation, and enzymatic digestion steps, which require significant analyst hands-on time while limiting result turnaround. Such complexity can also render nontrivial variations across analysts and laboratories. We describe the development of a fully automated peptide mapping procedure with a high-throughput robotic liquid handling system to improve sample handling capability and outcome reproducibility while saving analyst hands-on time. The automated procedure is completely hands-free, and setup requires the analyst only to prenormalize the sample concentrations and load buffers and reagents at their designated positions on the robotic deck. The robotic liquid handler performs all the subsequent preparation steps and stores the digested samples on a chiller unit to await retrieval. The convenience and flexibility provided by this automated peptide mapping method provides substantial benefits over manual sample preparation protocols. The optimized, automated procedure showed good reproducibility and results that were comparable to those of the manual procedure with respect to sequence coverage, digestion completeness, and quantification of posttranslational modifications. With this increased throughput, coupled with fast MAM analysis, more comprehensive characterization can be achieved.

scholarly journals High-throughput genotyping of single nucleotide polymorphisms with rolling circle amplification

BMC Genomics ◽  
2001 ◽  
Vol 2 (1) ◽  
Author(s):  
A Fawad Faruqi ◽  
Seiyu Hosono ◽  
Mark D Driscoll ◽  
Frank B Dean ◽  
Osama Alsmadi ◽  
...  
Keyword(s):  
Single Nucleotide Polymorphisms ◽  
High Throughput ◽  
Rolling Circle Amplification ◽  
Nucleotide Polymorphisms ◽  
Single Nucleotide ◽  
Rolling Circle

scholarly journals ENDO-Pore: High-throughput linked-end mapping of single DNA cleavage events using nanopore sequencing

2021 ◽  
Author(s):  
Oscar E Torres Montaguth ◽  
Stephen J Cross ◽  
Kincaid W.A. Ingram ◽  
Laura Lee ◽  
Fiona M Diffin ◽  
...  
Keyword(s):  
High Throughput ◽  
Single Molecule ◽  
Dna Cleavage ◽  
Cleavage Site ◽  
Rolling Circle Amplification ◽  
Time Resolved ◽  
Rolling Circle ◽  
Precise Position ◽  
And Function

Mapping the precise position of DNA cleavage events plays a key role in determining the mechanism and function of endonucleases. ENDO-Pore is a high-throughput nanopore-based method that allows the time resolved mapping single molecule DNA cleavage events in vitro. Following linearisation of a circular DNA substrate by the endonuclease, a resistance cassette is ligated recording the position of the cleavage event. A library of single cleavage events is constructed and subjected to rolling circle amplification to generate concatemers. These are sequenced and used to produce accurate consensus sequences. To identify the cleavage site(s), we developed CSI (Cleavage Site Investigator). CSI recognizes the ends of the cassette ligated into the cleaved substrate and triangulates the position of the dsDNA break. We firstly benchmarked ENDO-Pore using Type II restriction endonucleases. Secondly, we analysed the effect of crRNA length on the cleavage pattern of CRISPR Cas12a. Finally, we mapped the time-resolved DNA cleavage by the Type ISP restriction endonuclease LlaGI that introduces random double-strand breaks into its DNA substrates.

scholarly journals Automated High Throughput Purification of BigDye™ Terminator Fluorescent DNA Sequencing Reactions Using Wizard™ MagneSil™ Paramagnetic Particles

2002 ◽  
Vol 7 (5) ◽  
pp. 75-79
Author(s):  
Paul Otto ◽  
Brad Larson ◽  
Steve Krueger
Keyword(s):  
Sequence Analysis ◽  
Dna Sequencing ◽  
High Throughput ◽  
Liquid Handling ◽  
Robotic Platforms

We describe a reagent system and robotic methods for the purification of BigDye™ Terminator sequencing reactions prior to automated fluorescent sequence analysis. The methods use MagneSil™ paramagnetic particles to isolate sequencing extension products from unincorporated dye-labeled terminators and exchanges sequencer loading solution for reaction buffer. Processed samples give usable data that is greater than 98% accurate from primer plus 5–15 bases to over 700 bases. Typical Phred greater than 20 quality scores range from 600 to over 700 bases. This process has been adapted to a number of liquid handling robotic platforms in both 96- and 384-well formats. One method using a single POD Beckman Biomek® FX can process up to four plates in approximately 40 minutes.

scholarly journals Assay Miniaturization for Ultra-High Throughput Screening of Combinatorial and Discrete Compound Libraries: A 9600-Well (0.2 Microliter) Assay System

1998 ◽  
Vol 3 (1) ◽  
pp. 55-62 ◽  
Author(s):  
Kevin R. Oldenburg ◽  
Ji-Hu Zhang ◽  
Tongming Chen ◽  
Anthony Maffia ◽  
Karl F. Blom ◽  
...  
Keyword(s):  
High Throughput ◽  
Combinatorial Chemistry ◽  
Therapeutic Target ◽  
High Throughput Screening ◽  
Detection System ◽  
Screening System ◽  
Handling System ◽  
Liquid Handling ◽  
Compound Libraries ◽  
Plate Design

Combinatorial chemistry has opened a new realm of chemical entities in the search for novel therapeutics. Combinatorial chemistry is currently adding hundreds of thousands of compounds to similar numbers available from years of synthesis by medicinal chemistry. It is not unreasonable to expect that over the next several years, nearly a million compounds will be available for screening against each therapeutic target. The number of potential targets will also be increasing with the advances in genomics. With the increasing number of compounds to be screened against an increasing number of targets, it is becoming increasingly difficult and costly to obtain the required amounts of key biological material needed to screen these compounds. One obvious solution is to miniaturize the assays so that the biological reagent supply doesn't need to increase. To this end, we have developed an ultra-high throughput screening system comprised of a new plate design (9600-well), detection system, and liquid handling system. This new format is capable of performing assays in as little as 0.2 Al. The results obtained from this system compare favorably to those obtained in the standard 96-well format.

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