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Nanodrop nd 100

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The NanoDrop ND-100 is a compact and versatile spectrophotometer designed for the quantification and analysis of small volume samples. It utilizes a patented sample retention system to measure the absorbance of samples as small as 1 microliter, making it suitable for a wide range of applications, including DNA, RNA, and protein quantification.

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Lab products found in correlation

2100 bioanalyzer, by Agilent Technologies (2 mentions) Rneasy kit, by Qiagen (2 mentions) Amaxa nucleofector, by Lonza (2 mentions) Mikro s, by Sartorius (1 mentions) Tri reagent, by Thermo Fisher Scientific (1 mentions) Sybr green 1 master mix, by Roche (1 mentions) Mircury lna universal cdna synthesis kit 2, by Qiagen (1 mentions) Mirvana mirna isolation kit, by Thermo Fisher Scientific (1 mentions) Quantitec reverse transcription kit, by Qiagen (1 mentions) Lightcycler 480, by Roche (1 mentions)

4 protocols using nanodrop nd 100

1

Remodeller Depletion in ES Cells

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We used the pHYPER shRNA vector for remodeller depletion in ES cells, as previously described28 (link). shRNA design was performed using DESIR software (http://biodev.extra.cea.fr/DSIR/DSIR.html). Below are listed the shRNA selected for each remodeller. The sense strand sequence is given; the rest of the shRNA sequence is as described28 (link).
Each shRNA was transfected in its corresponding tagged ES cell line, in order to follow remodeller depletion by Western blotting using monoclonal antibodies anti-FLAG (M2, Sigma F1804), or anti-HA (H7, Sigma H3663) epitopes (Extended Data Fig. 6), in comparison with the signal obtained with a control antibody anti-GAPDH (Abcam ab9485). The pHYPER shRNA vectors were transfected in ES cell by electroporation, using an Amaxa nucleofector (Lonza). 24 h after transfection, puromycin (2 μg/ml) selection was applied for an additional 48 h period, before cell collection and RNA preparation, except for Chd4, for which cells were collected after 30 h of selection. Total RNA was extracted using an RNeasy Kit (Qiagen). Total RNA yield was determined using a NanoDrop ND-100 (Labtech). Total RNA profiles were recorded using a Bioanalyzer 2100 (Agilent). For each remodeller, RNA was prepared from three independent transfection experiments, and processed for transcriptome analysis.
de Dieuleveult M., Yen K., Hmitou I., Depaux A., Boussouar F., Dargham D.B., Jounier S., Humbertclaude H., Ribierre F., Baulard C., Farrell N.P., Park B., Keime C., Carrière L., Berlivet S., Gut M., Gut I., Werner M., Deleuze J.F., Olaso R., Aude J.C., Chantalat S., Pugh B.F, & Gérard M. (2016). Genome-wide nucleosome specificity and function of chromatin remodellers in ES cells. Nature, 530(7588), 113-116.
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2

RNA Extraction from Tendon Tissue

Cited 1 time
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Tendon was pulverising into a powder with a dismembranator (Mikro-S; Sartorius, Melsungen, Germany) following freezing in liquid nitrogen. Immediately, 20 volumes of Tri Reagent (Ambion) was added to the powdered tendon tissue and the RNA extracted and purified as described by Peffers et al. [25 (link)] (2013). RNA was quantified by using a Nanodrop ND-100 spectrophotometer (Labtech, Uckfield, East Sussex, UK) and assessed for purity by UV absorbance measurements at 260 and 280 nm.
Peffers M.J., Fang Y., Cheung K., Wei T.K., Clegg P.D, & Birch H.L. (2015). Transcriptome analysis of ageing in uninjured human Achilles tendon. Arthritis Research & Therapy, 17(1), 33.
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3

Quantitative RT-PCR for mRNA and miRNA

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Total and small RNAs were isolated using a mirVana miRNA Isolation Kit (Invitrogen) according to the manufacturer’s instructions and RNA eluted using RNase-free water. Both concentration and purity were measured by a spectrophotometer (NanoDrop ND-100, Labtech). The total RNA samples (100 ng) were reverse transcribed with a Quantitec Reverse Transcription Kit (Qiagen). Real-time PCR was performed using SYBR Green I Master Mix (Roche) and specific primers for mouse genes (Qiagen) in a LightCycler 480 (Roche). Duplicate CTs were averaged, and results analyzed by the 2−ΔΔCT method using 18S or Actb as a housekeeper gene. For miRNA detection, 5 ng/μL was used from each small RNA template, and cDNA synthesized using the miRCURY LNA Universal cDNA Synthesis kit II (Qiagen). Real-time PCR for miR-21-5p, -155, and -706 was performed using SYBR Green I Master Mix in a LightCycler 480. Duplicates of CTs were averaged, and the relative quantities of miRNAs calculated using the 2−ΔΔCT method and normalized to several artificial spike-ins as controls for extracellular miRNAs.
Zeboudj L., Sideris-Lampretsas G., Silva R., Al-Mudaris S., Picco F., Fox S., Chambers D, & Malcangio M. (2023). Silencing miR-21-5p in sensory neurons reverses neuropathic allodynia via activation of TGF-β–related pathway in macrophages. The Journal of Clinical Investigation, 133(11), e164472.
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4

Remodeller Depletion in ES Cells

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We used the pHYPER shRNA vector for remodeller depletion in ES cells, as previously described28 (link). shRNA design was performed using DESIR software (http://biodev.extra.cea.fr/DSIR/DSIR.html). Below are listed the shRNA selected for each remodeller. The sense strand sequence is given; the rest of the shRNA sequence is as described28 (link).
Each shRNA was transfected in its corresponding tagged ES cell line, in order to follow remodeller depletion by Western blotting using monoclonal antibodies anti-FLAG (M2, Sigma F1804), or anti-HA (H7, Sigma H3663) epitopes (Extended Data Fig. 6), in comparison with the signal obtained with a control antibody anti-GAPDH (Abcam ab9485). The pHYPER shRNA vectors were transfected in ES cell by electroporation, using an Amaxa nucleofector (Lonza). 24 h after transfection, puromycin (2 μg/ml) selection was applied for an additional 48 h period, before cell collection and RNA preparation, except for Chd4, for which cells were collected after 30 h of selection. Total RNA was extracted using an RNeasy Kit (Qiagen). Total RNA yield was determined using a NanoDrop ND-100 (Labtech). Total RNA profiles were recorded using a Bioanalyzer 2100 (Agilent). For each remodeller, RNA was prepared from three independent transfection experiments, and processed for transcriptome analysis.
de Dieuleveult M., Yen K., Hmitou I., Depaux A., Boussouar F., Dargham D.B., Jounier S., Humbertclaude H., Ribierre F., Baulard C., Farrell N.P., Park B., Keime C., Carrière L., Berlivet S., Gut M., Gut I., Werner M., Deleuze J.F., Olaso R., Aude J.C., Chantalat S., Pugh B.F, & Gérard M. (2016). Genome-wide nucleosome specificity and function of chromatin remodellers in ES cells. Nature, 530(7588), 113-116.
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