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Epmotion 5075 robot

Manufactured by Eppendorf
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Sourced in Germany

The EpMotion 5075 is a compact, automated liquid handling robot designed for precise and reproducible liquid transfers. It features a high-performance pipetting system with disposable tips and can handle a wide range of sample volumes. The EpMotion 5075 is suitable for a variety of laboratory applications that require accurate liquid handling.

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Lab products found in correlation

Hiseq 2000, by Illumina (2 mentions) Qubit fluorometry, by Thermo Fisher Scientific (2 mentions) Hiseq 4000, by Illumina (2 mentions) Hiseq 2000 platform, by Illumina (2 mentions) Qubit 2.0 fluorometer, by Thermo Fisher Scientific (2 mentions) Powersoil dna kit, by Eppendorf (2 mentions) 515f 806r primers, by Illumina (2 mentions) Iscript cdna synthesis kit, by Bio-Rad (1 mentions) Sybr select master mix, by Thermo Fisher Scientific (1 mentions) Quantstudio 12 k flex real timer, by Thermo Fisher Scientific (1 mentions) Synergy ht plate reader, by Agilent Technologies (1 mentions) Viral toxglo, by Promega (1 mentions) Truseq stranded total rna sample prep kit, by Illumina (1 mentions) Bioanalyzer dna 1000 chip, by Agilent Technologies (1 mentions) Ribo zero gold, by Illumina (1 mentions) Ampure bead, by Beckman Coulter (1 mentions) Sample prep kit, by Illumina (1 mentions) Agilent bioanalyzer dna 1000 chip, by Agilent Technologies (1 mentions) Truseq rna sample prep kit v2, by Illumina (1 mentions) Truseq sbs sequencing kit version 3, by Illumina (1 mentions)

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EpMotion 5075 (71 citations)

13 protocols using epmotion 5075 robot

1

Quantitative PCR Analysis of Heterologous and Native Genes

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cDNA was synthetized with the iScript™ cDNA Synthesis kit (Bio-Rad, Hercules, CA, USA), following the manufacturer’s instructions. For qPCR, a set of primers were designed for specific target cDNA. The set of selected genes comprised CRL1 (heterologous gene); AOX1, the alcohol oxidase 1 native gene; and MIT1, which codifies a key transcription factor of the methanol-induced transcription. Furthermore, KAR2 and HAC1, two genes involved in the unfolded protein response (UPR), were also analyzed.
For qPCR, reactions were done with SYBR™ Select Master Mix (Thermo Scientific™ Waltham, MA, US). Additionally, and as suggested by the manufacturer, to assure the maximum accuracy the reaction mixes were made by EpMotion® 5075 robot (Eppendorf, Germany).
The amplification program was run on a QuantStudio 12 K Flex Real-Timer from Thermo Scientific™ (Waltham, MA, US), following the manufacturer’s instructions. The annealing extension temperature was set at 57.4 °C. Relative transcript levels (RTLs) were determined by using MTH1 as a housekeeping gene as it shows basal expression across the conditions tested.
Garrigós-Martínez J., Nieto-Taype M.A., Gasset-Franch A., Montesinos-Seguí J.L., Garcia-Ortega X, & Valero F. (2019). Specific growth rate governs AOX1 gene expression, affecting the production kinetics of Pichia pastoris (Komagataella phaffii) PAOX1-driven recombinant producer strains with different target gene dosage. Microbial Cell Factories, 18, 187.
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2

Anti-Ebola Virus Drug Screening

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All experiments using infectious EBOV were performed under biosafety level 4 (BSL-4) conditions at the Galveston National Laboratory. Vero E6 cells (1 × 104 cells/well) were plated in 96-well plates (black clear bottom, Costar) overnight, and the next day compounds (azacitidine, cycloheximide, emetine, gedunin, or mycophenolic acid) were added at 0.4, 2, 10, and 50 μM using an epMotion 5075 robot (Eppendorf). One-hour post-treatment, EBOV-GFP3 was added at an MOI of 0.3 (40 μL) to media containing compound (60 μL) and left on the cells for the course of the experiment. Three days post-infection the mean fluorescence intensity (MFI) was measured using an EnVision plate reader. To assess cell viability, 1 × 104 cells/well were plated in 96-well plates (white polystyrene, Costar) overnight and treated with compound as above. Cell viability was assessed 5 days after compound treatment using Viral ToxGlo (Promega), and ATP content was determined by reading luminescence using a BioTek Synergy HT plate reader.
Edwards M.R., Pietzsch C., Vausselin T., Shaw M.L., Bukreyev A, & Basler C.F. (2015). High-Throughput Minigenome System for Identifying Small-Molecule Inhibitors of Ebola Virus Replication. ACS infectious diseases, 1(8), 380-387.
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3

Illumina Stranded Total RNA Sequencing

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RNA libraries were prepared according to the manufacturer’s instructions for the TruSeq Stranded Total RNA Sample Prep Kit (Illumina, San Diego, CA). The liquid handling Eppendorf (Hamburg, GER) EpMotion 5075 robot was employed for all AMPure bead clean up. Reverse transcription, A-tailing reaction, and adaptor ligation steps were performed manually. Briefly, 1 μg of total RNA was used as input for ribosomal depletion by RiboZero Gold™ (Illumina) to remove both cytoplasmic and mitochondrial rRNA. First strand cDNA synthesis was performed using SuperScript III reverse transcriptase, Actinomycin D, and random primers. Second strand cDNA was synthesized using dUTP. The stranded cDNA ends were A-tailed and ligated with index adaptors for multiplex sequencing. The adapter-modified DNA fragments were enriched by 15 cycles of PCR using primers included in the Illumina Sample Prep Kit. The concentration and size distribution of the libraries were determined on an Agilent Bioanalyzer DNA 1000 chip (Santa Clara, CA). A final quantification, using Qubit fluorometry (Invitrogen, Carlsbad, CA), was done to confirm sample concentration. The libraries were sequenced as 101 paired end reads on an Illumina HiSeq 2000.
Tricoli J.V., Boardman L.A., Patidar R., Sindiri S., Jang J.S., Walsh W.D., McGregor PM I.I.I., Camalier C.E., Mehaffey M.G., Furman W.L., Bahrami A., Williams P.M., Lih C.J., Conley B.A, & Khan J. (2017). A Mutational Comparison of Adult and Adolescent and Young Adult (AYA) Colon Cancer. Cancer, 124(5), 1070-1082.
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4

TruSeq RNA Library Prep on Eppendorf EpMotion

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RNA libraries were prepared according to the manufacturer’s instructions for the TruSeq RNA Sample Prep Kit v2 (Illumina). The liquid-handling EpMotion 5075 robot (Eppendorf) was used for TruSeq library construction. All AMPure bead (Beckman Coulter) clean up, mRNA isolation, end repair, and A-tailing reactions were completed on the 5075 robot. Reverse transcription and adaptor ligation steps were performed manually. The adapter-modified DNA fragments were then enriched by 12 cycles of polymerase chain reaction (PCR) using primers included in the Illumina Sample Prep Kit. Concentration and size distribution of the libraries were determined on an Agilent Bioanalyzer DNA 1000 chip. Sample concentration was confirmed using Qubit fluorometry (Invitrogen). Libraries were loaded onto paired-end flow cells at concentrations of 8 to 10 pM to generate cluster densities of 700,000/mm2 following Illumina’s standard protocol using the Illumina cBot and cBot Paired-End Cluster Kit version 3. The flow cells were sequenced as 51×2 paired-end reads on an Illumina HiSeq 2000 using TruSeq SBS Sequencing Kit version 3 and HCS v2.0.12 data collection software. Base-calling was performed using Illumina’s RTA version 1.17.21.3.
Shih Y.H., Zhang Y., Ding Y., Ross C.A., Li H., Olson T.M, & Xu X. (2015). The Cardiac Transcriptome and Dilated Cardiomyopathy Genes in Zebrafish. Circulation. Cardiovascular genetics, 8(2), 261-269.
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5

Quantitative Gene Expression Analysis

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Gene expression was measured as already described [15 (link)]. Briefly, tumor samples, with more than 80% of tumor cells, were homogenized in RNA lysis buffer in ice with an Ultra-Turrax (IKA, Staufen, Germany), and RNA was purified using the Maxwell 16 LEV SimplyRNA Cells kit (Promega, Madison, WI, USA). In all the samples by real time-PCR, the % of murine DNA contamination was established using primers specifically designed to distinguish human from murine actin and only samples with more than 85% human DNA were processed. Retro-transcription to cDNA was done using the High-Capacity cDNA Reverse Transcription kit (Thermo Fisher Scientific, Waltham, MA, USA). Genes selected were DNA pol β, ERCC1 and XPF. Optimal primer pairs (Table S3) were chosen, spanning splice junctions, using Primer3 Input software (Primer3 Input. http://primer3.ut.ee/) and the human specificity was verified by detecting single-band amplicons of the PCR products. Absolute copy numbers of mRNA were determined by real time-PCR (ABI-7900, Applied Biosystems, Foster City, CA, USA) with the SYBR Green technique (Promega), using an EP Motion 5075 robot (Eppendorf, Hamburg, Germany). Standard curves for each gene were included for absolute quantification of mRNA.
Real time-PCR data were normalized using the geometric mean of two housekeeping genes, actin (ACTB) and ciclophillin (CYPA).
Guffanti F., Alvisi M.F., Caiola E., Ricci F., De Maglie M., Soldati S., Ganzinelli M., Decio A., Giavazzi R., Rulli E, & Damia G. (2020). Impact of ERCC1, XPF and DNA Polymerase β Expression on Platinum Response in Patient-Derived Ovarian Cancer Xenografts. Cancers, 12(9), 2398.
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6

Dust Metagenomic Analysis Protocol

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Dust was collected at each site using a vacuum fitted with Dustream collectors (Indoor Biotechnologies, Charlottesville, VA). Collector filters (40-μm-pore mesh) containing the vacuumed dust were placed into sterile Nasco Whirl-Paks bags (Nasco, Fort Atkinson, WI) and stored under dark conditions at room temperature during collections. Collection took place from 18 July to 21 November 2016. Each sample was homogenized, and 0.25 g of dust was aliquoted into sterile 2-ml tubes and stored at −80°C until DNA extraction.
The DNA from dust aliquots was extracted using the MoBio PowerLyzer PowerSoil DNA isolation kit (MoBio, Carlsbad, CA, USA) protocol. To perform a metagenomic analysis, 1 ng of gDNA from each sample was prepared using the Illumina Nextera XT DNA library prep kit, along with the corresponding Illumina index kits v2 set A and set B, following the manufacturer’s instructions through the amplification step. Amplified products were purified with a modified bead-based DNA cleanup protocol using Mag-Bind RxnPure Plus by Omega Bio-Tek (Norcross, GA), quantified using the Quant-iT double-stranded DNA (dsDNA) assay kit, and pooled with equal concentrations of product using an Eppendorf (Hamburg, DE) epMotion 5075 robot. Libraries were sequenced on an Illumina HiSeq 4000 with 150-bp paired-end reads (insert size ranged from 250 to 1,000 bp).
Fahimipour A.K., Ben Mamaar S., McFarland A.G., Blaustein R.A., Chen J., Glawe A.J., Kline J., Green J.L., Halden R.U., Van Den Wymelenberg K., Huttenhower C, & Hartmann E.M. (2018). Antimicrobial Chemicals Associate with Microbial Function and Antibiotic Resistance Indoors. mSystems, 3(6), e00200-18.
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7

Molecular Profiling of cDDP-Treated Tumors

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Total mRNA was extracted from snap-frozen tissues by using Maxwell 16 LEV SimplyRNA (Promega), according to manufacturer protocols. Control/no treated (CTR) and cDDP-treated re-growing tumors (after the second cDDP cycle) were snap-frozen at the time of sacrifice, when tumor weights were about 400–1000 mg. The RT2 Profiler PCR Arrays (Qiagen) are designed to analyze a panel of genes related to EMT and CSCs. Each array contains a panel of 4×96 primer sets for a thoroughly researched set of 84 genes, plus five housekeeping genes, three mRNA retrotranscription and three PCR quality controls. For each plate two control/no-treated and two cDDP-treated samples of the same xenograft were included, except for MNHOC212 and MNHOC230 for which only control no-treated samples were analyzed. The assay includes a specific step of mRNA retro-transcription, using the RT2 First Strand Kit. The plate was filled by an EP Motion 5075 robot (Eppendorf), so an excess of volume was used. Reactions were done on a 7900HT Sequence Detection System (Applied Biosystems).
Ricci F., Fratelli M., Guffanti F., Porcu L., Spriano F., Dell’Anna T., Fruscio R, & Damia G. (2016). Patient-derived ovarian cancer xenografts re-growing after a cisplatinum treatment are less responsive to a second drug re-challenge: a new experimental setting to study response to therapy. Oncotarget, 8(5), 7441-7451.
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8

Whole Brain Transcriptome Profiling

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Whole brain RNA was extracted from the 600 µL homogenate in RLT buffer after an additional 30 s homogenization following the Qiagen RNeasy Mini Kit protocol, including a DNase (Qiagen) treatment to remove genomic DNA. RNA quantities were determined for each sample using a Qubit RNA HS Assay Kit (Invitrogen, Carlsbad, CA). High RNA integrity for all samples was confirmed with Bioanalyzer 2100 RNA Pico chips (Agilent, Santa Clara, CA) prior to library preparation.
RNAseq libraries were constructed and sequenced by the W.M. Keck Center for Comparative and Functional Genomics at the Roy J. Carver Biotechnology Center (University of Illinois at Urbana-Champaign). Libraries were constructed from 500 ng RNA per sample using the TruSeq Stranded mRNA HT kit (Illumina, San Diego, CA) on an ePMotion 5075 robot (Eppendorf, Hamburg, Germany). Libraries were uniquely barcoded, quantified, and pooled for sequencing across 6 lanes with 100 nt single-end sequencing on the Illumina HiSeq 4000.
Jones B.M., Rao V.D., Gernat T., Jagla T., Cash-Ahmed A.C., Rubin B.E., Comi T.J., Bhogale S., Husain S.S., Blatti C., Middendorf M., Sinha S., Chandrasekaran S, & Robinson G.E. (2020). Individual differences in honey bee behavior enabled by plasticity in brain gene regulatory networks. eLife, 9, e62850.
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9

Fecal Microbiome DNA Extraction and Sequencing

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Genomic DNA was extracted from 0.25 g of fecal material from each sample using the Earth Microbiome DNA extraction protocol28 . Briefly, DNA was extracted using the 96-well format MoBio Powersoil DNA kit on an EpMotion 5075 robot with vacuum (Eppendorf, Hamburg, Germany). DNA was quantified with the Qubit 2.0 fluorometer (Invitrogen, Carlsbad, CA) according to the manufacturer’s instructions.
PCR amplification and library preparation were performed similarly to the protocol described by Caporaso et al.29 (link). 515F/806R Illumina primers with unique reverse primer barcodes were used to target the V4 region of the 16S rRNA gene. Samples were amplified in triplicate and cleaned using the MO BIO 69 htp PCR cleanup kit. Each PCR reaction included 1X PCR buffer, 10 μM each forward and reverse primer, 200 μM dNTPs, 1 U/ml Taq polymerase, 15 ng template DNA, and PCR grade water, with a total reaction volume of 25 μL. Reactions were kept at 94°C for 3 minutes for denaturation to occur. Amplification was performed by 25 cycles of 94 °C for 45s, 58 °C for 60s, and 72 °C for 90s. The V4 amplicons were sequenced on the Illumina HiSeq 2000 platform, yielding single end, 100 base pair reads. Sequencing and quality assessment were performed at the Yale Center for Genome Analysis.
Halfvarson J., Brislawn C.J., Lamendella R., Vázquez-Baeza Y., Walters W.A., Bramer L.M., D’Amato M., Bonfiglio F., McDonald D., Gonzalez A., McClure E.E., Dunklebarger M.F., Knight R, & Jansson J.K. (2017). Dynamics of the human gut microbiome in Inflammatory Bowel Disease. Nature microbiology, 2, 17004.
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10

Fecal Microbiome DNA Extraction and Sequencing

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Genomic DNA was extracted from 0.25 g of fecal material from each sample using the Earth Microbiome DNA extraction protocol28 . Briefly, DNA was extracted using the 96-well format MoBio Powersoil DNA kit on an EpMotion 5075 robot with vacuum (Eppendorf, Hamburg, Germany). DNA was quantified with the Qubit 2.0 fluorometer (Invitrogen, Carlsbad, CA) according to the manufacturer’s instructions.
PCR amplification and library preparation were performed similarly to the protocol described by Caporaso et al.29 (link). 515F/806R Illumina primers with unique reverse primer barcodes were used to target the V4 region of the 16S rRNA gene. Samples were amplified in triplicate and cleaned using the MO BIO 69 htp PCR cleanup kit. Each PCR reaction included 1X PCR buffer, 10 μM each forward and reverse primer, 200 μM dNTPs, 1 U/ml Taq polymerase, 15 ng template DNA, and PCR grade water, with a total reaction volume of 25 μL. Reactions were kept at 94°C for 3 minutes for denaturation to occur. Amplification was performed by 25 cycles of 94 °C for 45s, 58 °C for 60s, and 72 °C for 90s. The V4 amplicons were sequenced on the Illumina HiSeq 2000 platform, yielding single end, 100 base pair reads. Sequencing and quality assessment were performed at the Yale Center for Genome Analysis.
Halfvarson J., Brislawn C.J., Lamendella R., Vázquez-Baeza Y., Walters W.A., Bramer L.M., D’Amato M., Bonfiglio F., McDonald D., Gonzalez A., McClure E.E., Dunklebarger M.F., Knight R, & Jansson J.K. (2017). Dynamics of the human gut microbiome in Inflammatory Bowel Disease. Nature microbiology, 2, 17004.
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