Anchored multiplex amplification on a microelectronic chip array

Westin, Lorelei; Xu, Xing; Miller, Carolyn T.; Wang, Ling; Edman, Carl F.; Nerenberg, Michael

doi:10.1038/72658

Cited by 106 publications

(63 citation statements)

References 23 publications

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“…Figure 2 shows the results of a titration experiment with various amounts of Chlamydia template input. Noncleavable primers, which have a single base mutation in the recognition site for BsoBI, were used as a control for background binding of hybridized template and to ensure that amplification was occurring on the microchip surface (22). Template input levels of 1,000 copies of DNA could be routinely detected using anchored SDA in multiple experiments.…”

Section: Resultsmentioning

confidence: 99%

“…3a, only S. aureus samples that have been characterized as methicillin resistant showed any positive signal. An anchored SDA system from the human factor V gene (22) was used as a control to show that amplification was functional in the methicillin-sensitive amplification reaction. These results demonstrate the specificity and accuracy of anchored SDA.…”

Section: Resultsmentioning

confidence: 99%

“…However, most microchip array systems require large amounts of template DNA, or targets from multiplex amplification, in order to detect DNA or RNA at fairly low levels (2, 13). We recently described an assay on an electronic microchip array which was capable of multiplex amplification of 10 targets simultaneously, with little decrease in amplification efficiency (22). Anchored SDA on microelectronic chips encompasses amplification on the surface of the array, requiring very small amounts of input DNA.…”

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confidence: 99%

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Antimicrobial Resistance and Bacterial Identification Utilizing a Microelectronic Chip Array

Westin¹,

Miller²,

Vollmer³

et al. 2001

J Clin Microbiol

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Species-specific bacterial identification of clinical specimens is often limited to a few species due to the difficulty of performing multiplex reactions. In addition, discrimination of amplicons is time-consuming and laborious, consisting of gel electrophoresis, probe hybridization, or sequencing technology. In order to simplify the process of bacterial identification, we combined anchored in situ amplification on a microelectronic chip array with discrimination and detection on the same platform. Here, we describe the simultaneous amplification and discrimination of six gene sequences which are representative of different bacterial identification assays: Escherichia coli gyrA, Salmonella gyrA, Campylobacter gyrA, E. coli parC, Staphylococcus mecA, and Chlamydia cryptic plasmid. The assay can detect both plasmid and transposon genes and can also discriminate strains carrying antibiotic resistance single-nucleotide polymorphism mutations. Finally, the assay is similarly capable of discriminating between bacterial species through reporter-specific discrimination and allele-specific amplification. Anchored strand displacement amplification allows multiplex amplification and complex genotype discrimination on the same platform. This assay simplifies the bacterial identification process greatly, allowing molecular biology techniques to be performed with minimal processing of samples and practical experience.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

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Antimicrobial Resistance and Bacterial Identification Utilizing a Microelectronic Chip Array

Westin¹,

Miller²,

Vollmer³

et al. 2001

J Clin Microbiol

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“…A multiplex SDA for up to ten targets was shown by Westin et al in a microarray chip with anchored primers [134]. Table 5 lists different strategies for the application of SDA in microsystems.…”

Section: Sda: Strand Displacement Amplificationmentioning

confidence: 99%

Isothermal Amplification and Quantification of Nucleic Acids and its Use in Microsystems

Tröger¹,

Niemann²,

Gärtig³

et al. 2015

J Nanomed Nanotechnol

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Nucleic acid amplification technologies (NAATs) offer the most sensitive tests in the clinical laboratory. These techniques are used as a powerful tool for screening and diagnosis of infectious diseases. Isothermal methods, as an alternative to polymerase chain reaction (PCR), require no thermocycling machine and can mostly be performed with reduced time, high throughput, and accurate and reliable results.However, current molecular diagnostic approaches generally need manual analysis by qualified and experienced personal which is a highly complex, time-consuming and labor-intensive task. Thus, the demand for simpler, miniaturized systems and assays for pathogen detection is steadily increasing. Microfluidic platforms and lab-on-a-chip devices have many advantages such as small sample volume, portability and rapid detection time and enable point-of-care diagnosis.In this article, we review several isothermal amplification methods and their implementation in microsystems in relation to quantification of nucleic acids. miniaturized systems can be diverse. The majority of publications are focussed on clinical diagnostics including foodborne organisms for environmental monitoring [10][11][12]. In case of an infectious disease, cultivation and phenotypic characterization are still the standard methods of microbiologists which need days up to weeks to obtain a diagnostic result. Hence, scientists started to make use of nucleic acid amplification methods to accelerate the analysis. The most widespread and well known technique for this purpose is the polymerase chain reaction (PCR) [10,13], which exploits the activity of the DNA polymerase. The method is based on a thermal cycling process consisting of repeated cycles of heating and cooling steps allowing for DNA melting, sequence specific primer binding and enzymatic amplification of the target nucleic acid. The quantitative assessment of target copies is possible by using a real-time PCR approach, where a concentration dependent signal (e.g. fluorescence) increases over time [14]. The latter enables the user to easily assess the pathogen load of patient samples [15][16][17][18]. Since the first example of a PCR on a miniaturized system in 1994 [19,20], numerous devices were presented, which integrate not just the amplification, but also sample preparation and detection steps [9,[21][22][23][24][25][26][27][28]. Just a few of those microsystems have reached the market so far, but some have shown a fascinating potential to replace the classical laboratory-oriented state-of-the art [29][30][31][32][33]. setups has been shown to obtain a similar result [34]. A smaller heat capacity allows for rapid changes in temperature beneficial for the PCR time as well as a higher parallelism of multiple genetic samples [34].However, a major drawback of the integration of PCR to microsystems is the necessity of sophisticated instrumentation. The reaction requires a thermal cycling instrumentation, space and considerable expertise [35]. Therefore, isothermal reactions for the ta...

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“…Initial applications are for SNP scoring or genotyping. Chips are being developed with 25, 100, 400, and 10,000 test sites for various applications such as infectious disease diagnostics, genomic research, genetic disease and cancer diagnostics, drug discovery, and gene expression Edman et al 2000;Radtkey et al 2000;Westin et al 2000).…”

Section: Mass Spectrometrymentioning

confidence: 99%

Automation for Genomics, Part Two: Sequencers, Microarrays, and Future Trends

Meldrum¹

2000

Genome Res.

116

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Automation for genomics has enabled a 43-fold increase in the total finished human genomic sequence in the world in the past four years. This is the second half of a two-part, noncomprehensive review that presents an overview of different types of automation equipment used in genome sequencing. The first part of the review, published in the previous issue, focused on automated procedures used to prepare DNA for sequencing or analysis. This second part of the review presents a look at available DNA sequencers and array technology and concludes with a look at future technologies. Alternate sequencing technologies including mass spectrometry, biochips, and single molecule analysis are included in this review.Sequencing data, essential for many of the medical breakthroughs of today and tomorrow, is currently accumulating at an exponential rate, largely because of advances in sequencing methods and laboratory automation. Instruments are available that can automate nearly every step in the large-scale sequencing process.The first article in this two-part review (Meldrum 2000) presented a description of automation designed for isolating DNA, cloning or amplifying DNA, preparing enzymatic sequencing reactions, and purifying DNA. Part one of the review also showed that the majority of genome centers use both in-house automation and commercial automation (Collins et al. 1998;Wendl et al. 1998;Mullikin and McMurragy 1999;Spurr et al. 1999; see http://www.genome.org for a supplementary table providing a snapshot view of automation used at a number of different genome centers).In this, the second part of the review, the focus is on automation after DNA preparation and sequencing reactions-on obtaining the sequence and its analysis. A discussion of future technologies for DNA sequencing projects is also included. (See the web sites referenced for photos or further details on the instruments presented.)

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Anchored multiplex amplification on a microelectronic chip array

Cited by 106 publications

References 23 publications

Antimicrobial Resistance and Bacterial Identification Utilizing a Microelectronic Chip Array

Antimicrobial Resistance and Bacterial Identification Utilizing a Microelectronic Chip Array

Isothermal Amplification and Quantification of Nucleic Acids and its Use in Microsystems

Automation for Genomics, Part Two: Sequencers, Microarrays, and Future Trends

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