The Genetic Modification Background_Adverse Effects of Genome Editing on Cellular Homeostasis
Ontology highlight
ABSTRACT:
INSTRUMENT(S): maXis 4G
ORGANISM(S): Streptococcus Pyogenes (ncbitaxon:1314) Staphylococcus Aureus (ncbitaxon:1280)
SUBMITTER: 
David Gonzalez
PROVIDER: MSV000090542 | MassIVE | Mon Oct 17 15:10:00 BST 2022
REPOSITORIES: MassIVE
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Project description:Samples for metabolomic analysis were prepared as follows. A 200 uL aliquot of bacterial cultures or control medium (prepared, as described in the above section) was mixed with 800 uL of 99.5 percent ethanol (Fisher Scientific) and subjected to 3 rounds of freeze thaw cycles: 10 minutes at -80C, 8 minutes at room temperature. Next, samples were spun down for 10 minutes at 16,000g, 4C to remove cell debris, and supernatants were transferred to new tubes. Ethanol supernatants were stored at -80C until further extraction. Each ethanol extraction was performed in two technical duplicates.
The solid phase extraction (SPE) of the ethanol extracts was performed with the Oasis HLB 96-well plate (30 mg Sorbent per Well, 30 um Particle Size) (Waters Corporation) set up with a vacuum pump manifold. Each well of the plate was washed with 700 uL of 100 percent HPLC-grade methanol and equilibrated with 700 uL of HPLC-grade water. Next, 400 uL of ethanol extracts were added to the wells and allowed to slowly elute. Wells were next washed with 800 uL of 5 percent methanol in water. Metabolites from bacterial cultures or control media were eluted with 200 uL of 100 percent methanol. Vacuum up to 20 psi was applied for the wells that did not elute within one hour. The collected eluted metabolites were stored at -20 until the HPLC-MS analysis. The HPLC-MS analysis of extracted metabolites was performed with the use of a Dionex UltiMate 3000 UHPLC system (Fisher Scientific) coupled to a Bruker impact HD quadrupole time-of-flight mass spectrometer (Bruker). The chromatographic separation was performed on a Kinetex C18 1.7 um, 100A UHPLC column (50 mm x 2.1 mm) (Phenomenex), held at 40C during analysis. 5 uL of each sample was injected. Mobile phase A was water, 0.1 percent v:v formic acid, and mobile phase B was acetonitrile, 0.1 percent v:v formic acid. The solvent gradient table was set as follows: 2 percent B, increased to 10 percent B over 0.2 min, then to 100 percent B at 12 min, held at 100 percent B for 1.5 min, decreased back to 2 percent B in 0.5 min, followed by washout cycle and equilibration; total analysis time of 15 min. The scanned m/z range was 80-2000 Th; capillary voltage 4,500 V; nebulizer gas pressure 2 bar, drying gas flow rate 9 l min-1 and temperature 200C. The full MS scan was followed by a tandem mass spectrometry fragmentation of the seven most abundant ions within the spectrum. For tandem mass spectrometry, the collision cell collision energy was set at 3 eV and the collision energy was stepped 50 percent, 75 percent, 150 percent, and 200 percent. The scan rate was 3 Hz. A HP-921 lock mass compound was infused during the analysis for post processing mass correction.
2022-10-17 | MSV000090542 | GNPS
Project description:MicroPoly(A)Pure kits (Ambion) were used to prepare mRNA from P. salmonis-infected and control Atlantic salmon head kidney. Head kidney mRNA samples were prepared from pooled (n = 10) infected or control kidney. Approximately 200 ng of infected or control head kidney mRNA was used as template in aRNA synthesis reactions. Two infected head kidney samples were pooled to yield 8.9 ug of aRNA, and a single control head kidney sample contained 8.4 ug of aRNA. Infected and control head kidney aRNA samples were visualized on agarose gels to check quality and quantity (data not shown). Except for the amounts of aRNA and reverse transcription (RT) primer used, head kidney target labeling reactions and microarray hybridizations were identical. Head kidney target syntheses used 2 ug of aRNA and 2 ug of random primers (Roche). RT primers were mixed with aRNA samples, and nuclease-free H2O (Invitrogen) was added to make 10 ul total volume. Samples were incubated at 70 degrees C for 4 min, then placed on ice. RT reactions included 50 mM Tris HCl pH 8.3, 75 mM KCl, 3 mM MgCl2, 10 mM DTT, 0.5 mM each dATP, dCTP, and dGTP, 0.2 mM dTTP, 0.05 mM Cy3-dUTP or Cy5-dUTP (Amersham), 40U RNAguard ribonuclease inhibitor (Amersham), and 400U Superscript II Rnase H- reverse transcriptase (Invitrogen). RT reactions were incubated at 42 degrees C for 2.5 h, followed by addition of 0.025U RNase A (Sigma) and 1.5U RNase H (New England BioLabs) and incubation at 37 degrees C for 30 min. Labeled targets were cleaned using the Qiaquick PCR purification kit (Qiagen) and precipitated, with 300 mM sodium acetate, 1 ul of 20 ug/ul glycogen (Invitrogen), and 2.5 volumes of 100% ethanol, at -20 degrees C overnight. Each labeled target was recovered by centrifugation (14,000 rpm, 4 degrees C, 1 h), washed in 70% ethanol, air-dried, and resuspended in 60 ul of hybridization buffer: 50% deionized formamide, 5X SSC, 0.1% SDS, 5 ul of 5 ug/ul oligo dT (5’T18[Q-N]3’), 5 ul of 2 ug/ul BSA (Pierce), and 2 ul of 10 ug/ul sonicated human placental DNA (Sigma). Targets were incubated at 96 degrees C for 3 min, and then at 65 degrees C until applied to microarrays. Microarrays were prepared for hybridization by washing 2 X 5 min in 0.1% SDS, washing 5 X 1 min in MilliQ H2O, immersing 3 min in 95 degrees C MilliQ H2O, and drying by centrifugation (2000 rpm, 5 min, in 50 ml conical tube). Microarray hybridizations were run in the dark under HybriSlips hybridization covers (Grace Biolabs) in slide hybridization chambers (Corning) submerged in a 45 degrees C water bath for 16 h. Coverslips were floated off in 45 degrees C (1X SSC, 0.2% SDS), and arrays were washed 1 X 10 min in 45 degrees C (1X SSC, 0.2% SDS), 3 X 4 min in room temperature (0.1X SSC, 0.1% SDS), and 4 X 4 min in room temperature 0.1X SSC. Slides were dried by centrifugation as before. Fluorescent images of hybridized arrays were acquired immediately at 10 um resolution using ScanArray Express (PerkinElmer). The Cy3 and Cy5 cyanine fluors were excited at 543 nm and 633 nm respectively, and the same laser power (90%) and photomultiplier tube (PMT) settings were used for all slides in a study (PMT 75 Cy3, PMT 63 or 64 Cy5 for head kidney study). Fluorescent intensity data were extracted from TIF images using Imagene 5.5 software (Biodiscovery). Keywords: other
2004-02-11 | GSE1031 | GEO
Project description:Transcript profiles of Postia placenta grown on media containing ball-milled aspen or ball-milled pine as the sole carbon source were analyzed. Array design was based on the DoE's Joint Genome Institute's gene models for P. placenta version 1. The research goal is to identtify genes essential for cellulose depolymerization. From a data set of 12,438 unique alleles, each NimbleGen (Madison, WI) array featured 10 unique 60mers per gene, all in triplicate. The dataset was manually annotated to include only the ‘best allelic model’ among CAZY-encoding genes. Total RNA was purified from medium containing ball-milled aspen or ball-milled pine as the sole carbon source. Three biological replicates per medium were used (6 separate arrays). RNA was converted to double-strand cDNA and labeled with the Cy3 fluorophore sample for hybridization to the Postia placenta MAD-698 whole genome expression array by Roche NimbleGen (Iceland). In brief, 10ug of total RNA was incubated with 1X first strand buffer, 10 mM DTT, 0.5mM dNTPs, 100 pM oligo T7 d(T)24 primer, and 200 units of SuperScript II (Invitrogen) for 60 min at 42°C. Second strand cDNA was synthesized by incubation with 1X second strand buffer, 0.2mM dNTPs, 0.07 units per ul DNA ligase (Invitrogen), 0.27 units per ul DNA polymerase I (Invitrogen), 0.013 units per ul RNase H (Invitrogen), at 16°C for 2 hours. Immediately following, 10 units T4 DNA polymerase (Invitrogen) was added for additional 5 minute incubation at 16°C. Double-stranded cDNA was treated with 27ng/ul of RNase A (EpiCenter Technologies) for 10 minutes at 37°C. Treated cDNA was purified using an equal volume of phenol:chloroform:isoamyl alcohol (Ambion), ethanol precipitated, washed with 80% ethanol, and resuspended in 20ul water. One ug of each cDNA sample was amplified and labeled with 1 unit per ul of Klenow Fragment (New England BioLabs) and 1 O.D unit of Cy3 fluorophore (TriLink Biotechnologies, Inc.) for 2 hours at 37°C. Array hybridization was carried out with 6ug of labeled cDNA suspended in NimbleGen hybridization solution for 17 hours at 42°C.
2011-06-02 | E-GEOD-29656 | biostudies-arrayexpress
Project description:MicroPoly(A)Pure kits (Ambion) were used to prepare mRNA from P. salmonis-infected and control Atlantic salmon macrophages. Infected and control (24 h incubation) macrophage mRNA samples were subjected to two rounds of amplification using the MessageAmp aRNA kit (Ambion) and manufacturer's instructions. Approximately 400 ng of macrophage mRNA was used as template in the first round of amplification, yielding 8.1 ug of infected macrophage aRNA and 7.8 ug of control macrophage aRNA. Approximately 1 ug of first-round macrophage aRNA was used as template in the second round of amplification, yielding 81.2 ug of infected macrophage aRNA and 83.3 ug of control macrophage aRNA. Infected and control macrophage aRNA samples were visualized on agarose gels to check quality and quantity. Except for the amounts of aRNA and reverse transcription (RT) primer used, macrophage target labeling reactions and microarray hybridizations were identical. Macrophage target syntheses used 5 ug of second-round aRNA and 5 ug of random primers (Roche). RT primers were mixed with aRNA samples, and nuclease-free H2O (Invitrogen) was added to make 10 ul total volume. Samples were incubated at 70 degrees C 4 min, then placed on ice. RT reactions included 50 mM Tris HCl pH 8.3, 75 mM KCl, 3 mM MgCl2, 10 mM DTT, 0.5 mM each dATP, dCTP, and dGTP, 0.2 mM dTTP, 0.05 mM Cy3-dUTP or Cy5-dUTP (Amersham), 40U RNAguard ribonuclease inhibitor (Amersham), and 400U Superscript II Rnase H- reverse transcriptase (Invitrogen). RT reactions were incubated at 42 degrees C for 2.5 h, followed by addition of 0.025U RNase A (Sigma) and 1.5U RNase H (New England BioLabs) and incubation at 37 degrees C for 30 min. Labeled targets were cleaned using the Qiaquick PCR purification kit (Qiagen) and precipitated, with 300 mM sodium acetate, 1 ul of 20 ug/ul glycogen (Invitrogen), and 2.5 volumes of 100% ethanol, at -20 degrees C overnight. Each labeled target was recovered by centrifugation (14,000 rpm, 4 degrees C, 1 h), washed in 70% ethanol, air-dried, and resuspended in 60 ul of hybridization buffer: 50% deionized formamide, 5X SSC, 0.1% SDS, 5 ul of 5 ug/ul oligo dT (5'T18[Q-N]3'), 5 ul of 2 ug/ul BSA (Pierce), and 2 ul of 10 ug/ul sonicated human placental DNA (Sigma). Targets were incubated at 96 degrees C for 3 min, and then at 65 degrees C until applied to microarrays. Microarrays were prepared for hybridization by washing 2 X 5 min in 0.1% SDS, washing 5 X 1 min in MilliQ H2O, immersing 3 min in 95 degrees C MilliQ H2O, and drying by centrifugation (2000 rpm, 5 min, in 50 ml conical tube). Microarray hybridizations were run in the dark under HybriSlips hybridization covers (Grace Biolabs) in slide hybridization chambers (Corning) submerged in a 45 degrees C water bath for 16 h. Coverslips were floated off in 45 degrees C (1X SSC, 0.2% SDS), and arrays were washed 1 X 10 min in 45 degrees C (1X SSC, 0.2% SDS), 3 X 4 min in room temperature (0.1X SSC, 0.1% SDS), and 4 X 4 min in room temperature 0.1X SSC. Slides were dried by centrifugation as before. Fluorescent images of hybridized arrays were acquired immediately at 10 um resolution using ScanArray Express (PerkinElmer). The Cy3 and Cy5 cyanine fluors were excited at 543 nm and 633 nm respectively, and the same laser power (90%) and photomultiplier tube (PMT) settings were used for all slides in a study (PMT 72 Cy3, PMT 63 or 64 Cy5 for macrophage study). Fluorescent intensity data were extracted from TIF images using Imagene 5.5 software (Biodiscovery). Keywords: other
2004-02-11 | GSE1012 | GEO
Project description:Female Sprague-Dawley rats were randomly separated into two groups: i) trained (Ex group) and ii) sedentary (Cont group). Animals from the Ex group were adapted to treadmill exercise for two consecutive weeks, involving a gradual increase in the running time till 60 min/day at 20 m/min, 0% grade, 5 days/week. This exercise program remained constant until the end of the 54 weeks. Mitochondria isolation from 10 animals (5 Ex group and 5 Cont group) was performed using the conventional methods of differential centrifugation. Mitochondrial protein extracts were reduced with dithiothreitol, alkylated with iodoacetamide and digested with trypsin using the FASP procedure (18). Briefly, each sample was solubilized in 4 % SDS, 0.1 M DTT, 0.1 M HEPES and incubated for 30 min at 60 C. Then, samples were mixed with 0.2 mL of a solution of 8 M Urea, 0.1 M HEPES, pH 8.5 (UH solution), loaded into the filtration devices (3k Microcon, Millipore), centrifuged at 14,000 g for 20 min, and washed twice with UH solution. After centrifugation, the concentrates were alkylated with 50 mM iodoacetamide in UH solution (dark, 25 C, 20 min), and washed twice with UH solution. The concentrate was diluted with 0.1 mL of 50 mM ammonium bicarbonate (ABC) and digested overnight (37 C) with the addition of 2 ug of trypsin diluted in 30 uL of 50 mM ABC. The resulting tryptic peptides were collected by centrifugation and the filter were washed twice with 50 uL of 50 mM ABC. Eluted peptides were acidified with 10 uL of formic acid and cleaned up on a homemade Empore C18 column (3M, St. Paul, MN, USA) (ref) for subsequent phospho-enrichment and/or analysis by LC-MS/MS. The dried protein digest was dissolved with 10 uL (3 % ACN, 0.1 % TFA), diluted 1:5 with loading buffer (1 M glycolic acid, 5 % TFA, 80 % ACN) and phosphopeptides were enriched on TiO2-beads as previously described (19). Briefly 5 mg of "Titansphere TiO2 5 um" were suspended in 100 uL of 100 % ACN, immobilized in a pipette-column and washed using 5 uL of loading buffer. Each sample was loaded in each pipette-column and then washed with 30 uL of washing buffer (1 % TFA, 80 % ACN). Phosphopeptides were eluted from the beads with 25 uL of 25 % ACN (v/v) containing 25 % NH4OH (m/v), acidified with 10 uL of 10 % TFA and vacuum- concentrated to approximately 4 uL. Samples were finally diluted to 10 uL with H2O + 0.1% formic acid prior to injection. The peptides mixtures were analyzed by online nanoflow liquid chromatography tandem mass spectrometry (nanoLC-MS/MS) on an EASY-nLC system (Proxeon Biosystems, Odense, Denmark) connected to the LTQ Orbitrap Velos instrument (Thermo Fisher Scientific, Bremen, Germany) through a nanoelectrospray ion source. Six microliters of the peptide mixture were auto-sampled directly onto the analytical HPLC column (120 mm x75 um I.D., 3 um particle size (Nikkyo Technos Co., Ltd.) with a 120-min gradient from 5 % to 50 % ACN in 0.1 % FA. The effluent from the HPLC column was directly electrosprayed into the mass spectrometer. The LTQ Orbitrap Velos instrument was operated in data-dependent acquisition mode to automatically switch between full scan MS and MS/MS acquisition. The MS survey scan was performed in the FT cell recording a window between 350 and 2000 m/z. The resolution was set to 60 000, and the automatic gain control was set to 1,000,000 ions with a maximal acquisition time of 400 ms. The 20 most intense peptide ions with charge states great than or equal to 2 were sequentially isolated to a target value of 5,000 and fragmented in the high-pressure linear ion trap by low-energy CID with normalized collision energy of 35% Q value of 0.25, and an activation time of 10 ms. Raw files were processed using Proteome Discoverer version 1.3 (Thermo Fisher Scientific, Bremen). Peak lists were searched using Mascot software version 2.3 (Matrix Science, UK) against a SwissProt database containing entries corresponding to Rattus norvegicus (version of July 2012), a list of common contaminants, and all the corresponding decoy entries. Trypsin was chosen as enzyme and a maximum of two miscleavages were allowed. Carbamidomethylation (C) was set as a fixed modification, whereas oxidation (M) and phosphorylation (STY) were used as variable modifications. Searches were performed using a peptide tolerance of 7 ppm, a product ion tolerance of 0.5 Da. Resulting data files were filtered for FDR less than 1 % at peptide level.
2014-02-14 | PXD000354 | Pride
Project description:Ammonium hydroxide and pyrrolidine eluents were dried (SpeedVac) and reconstituted in 25 ul sample loading buffer (0.1% TFA/2% acetonitrile). Eluent from phosphotyrosine immunoprecipitation was dried and reconstituted in 35 ul of the loading buffer. An LTQ Orbitrap XL (ThermoFisher) in-line with a Paradigm MS2 HPLC (Michrom bioresources) was employed for acquiring high-resolution MS and MS/MS data. Ten microliters of the phospho-enriched peptides were loaded onto a sample trap (Captrap, Bruker-Michrom) in-line with a nano-capillary column (Picofrit, 75 um i.d.x 15 um tip, New Objective) packed in-house with 10 cm of MAGIC AQ C18 reverse phase material (Michrom Bioresource). Two different gradient programs, one each for MOAC and phosphotyrosine immunoprecipitation samples, were used for peptide elution. For MOAC samples, a gradient of 5-40% buffer B (95% acetonitrile/1% acetic acid) in 135 min and 5 min wash with 100% buffer B followed by 30 min of re-equilibration with buffer A (2% acetonitrile/1% acetic acid) was used. For phosphotyrosine immunoprecipitation samples, which were a much less complex mixture of peptides, 5-40% gradient with buffer B was achieved in 75 min followed by 5 min wash with buffer B and 30 min re-equilibration. Flow rate was ~0.3 ul/min. Peptides were directly introduced into the mass spectrometer using a nano-spray source. Orbitrap was set to collect 1 MS scan between 400-2000 m/z (resolution of 30,000 @ 400 m/z) in orbitrap followed by data dependent CID spectra on top 9 ions in LTQ (normalized collision energy ~35%). Dynamic exclusion was set to 2 MS/MS acquisitions followed by exclusion of the same precursor ion for 2 min. Maximum ion injection times were set to 300 ms for MS and 100 ms for MS/MS. Automatic Gain Control (AGC) was set to 1xe6 for MS and 5000 for MS/MS. Charge state screening was enabled to discard +1 and unassigned charge states. Technical duplicate data for each of the MOAC elutions (ammonium hydroxide and pyrrolidine) and triplicate data for the phosphotyrosine immunoprecipitation samples were acquired. RAW files were converted to mzXML using msconvert and searched against the Swissprot Human protein database (2013-Jan release) appended with common proteomics contaminants and reverse sequences as decoys. Searches were performed with X!Tandem (version 2010.10.01.1) using the k-score plugin. For all searches the following search parameters were used: Parent monoisotopic mass error of 50 ppm; fragment ion error of 0.8 Daltons; allowing for up to 2 missed tryptic cleavages. Variable modifications were oxidation of Methionine (+15.9949@M), carbamidomethylation of Cysteine (+57.0214@C), and phosphorylation of Serine, Threonine, and Tyrosine (+79.9663@[STY]). The search results were then post-processed using PeptideProphet and ProteinProphet.
2014-01-09 | PXD000658 | Pride
Project description:We describe the use of conventional flow proteomics (200 uL/min) on an optimized system for the analysis of highly fractionated TMT labeled materials. We compare 20ug injections on column using NanoFreeProteomics (NFP) to 1uL injections on a convenentional nanoLC system
2020-02-25 | PXD016704 | Pride
Project description:Sample preparation: P-AMPK+AMP, P-AMPK+ATPγS, and AMPK+ATPγS protein samples were crosslinked in HBST buffer (25 mM HEPES pH 7.5 at room temperature, 200 mM NaCL, and 1 mM TCEP). Non-crosslinked negative controls were generated using the same procedure without the addition of DSSO. Reactions were initiated by spiking 1 uL of 75 mM DSSO (Thermo-Fisher) in DMSO into a 50 uL solution containing 10 uM protein and mixing. The final molar ratio of crosslinker:protein was 150:1. The reactions were incubated at 25°C for 45 minutes prior to being quenched by the addition of Tris pH 8.0 to a final concentration of 50 mM. Each crosslink reaction was done in triplicate and the reaction replicates were pooled after quenching. 10 μL samples of the pooled samples were loaded for SDS-PAGE analysis with Coomassie staining to confirm the presence of crosslinked protein. The remaining 140 μL of crosslinked and non-crosslinked samples were then acetone precipitated at -20°C overnight and the protein was pelleted by centrifugation at 16,000 rcf for 5 minutes at 4°C. After decanting, the pellets were dried for 15 minutes at room temperature before being resuspended in 12.5 uL of resuspension buffer (50 mM ammonium bicarbonate, 8M urea, pH 8.0). ProteaseMAX (Promega) was added to 0.02% and the solutions were mixed on an orbital shaker operating at 400 RPM for 5 minutes. After resuspension, 87.5 uL of digestion buffer (50 mM ammonium bicarbonate, pH 8.0) was added. Cysteines in the protein samples were reduced and akylated as follows. DTT was added to 5 mM final concentration and the protein solutions were incubated at 56°C in an orbital shaker operating at 400 RPM and for 20 minutes. After reduction, 2.7 uL of 550 mM iodoacetamide was added and the solutions were incubated in the dark at room temperature for 15 minutes. Reduced and alkylated protein solutions were digested using trypsin at a ratio of 1:200 (w/w trypsin:protein) and incubating at 37°C. After overnight incubation at 37°C, the samples were acidified by the addition of trifluoroacetic acid (TFA, Thermo-Fischer) to 1%. Samples were then frozen and stored at -20°C until analysis.
2021-07-08 | PXD019017 | Pride
Project description:Escherichia coli strain MG1655 was grown to mid-log phase in defined anaerobic media. One sample was treated with 100 µM CORM-3, the other sample was a control. The volume (175 ml), temperature (37oC) and stirring (200 rpm) were constant. After 15 min of exposure to CO-RM, samples were taken from treated and control cells, harvested into ice cold phenol ethanol (187 µl phenol, 3.56 ml ethanol) to stabilize RNA, and total RNA was purified using Qiagen’s RNeasy Mini kit as recommended by suppliers. RNA was quantified using a BioPhotometer (Eppendorf). Biological experiments were carried out four times, and a dye swap performed for each experiment, providing two technical repeats for each of the four biological repeats.
2008-12-22 | GSE12878 | GEO
Project description:We reported that the microRNA profiling of exosome isolated from 58 cerebrospinal fluid (CSF) of 38 medulloblastoma (MBL) patient with or without letomeningeal metastases (LM). CSF exosome was precipitated using Exo2DTM for RNA assay (EXOSOME plus, Suwon, South Korea) according to the manufacture’s protocol (Kim, et al. 2017). 2 ml of CSF were added to Exo2DTM reagent B following mixing by inverting, and incubated at room temperature for 30 min. The supernatant was discarded and the exosome containing pellet was resuspended in 100 ul of PBS to further RNA isolation. Total RNA was extracted from the CSF exosome fraction using a microRNA purification kit (Norgen Biotek corp., Thorold, Canada) according to the manufacture’s protocol. Briefly, samples were mixed with lysis buffer, and the lysate was added to absolute ethanol. The lysate was applied up to the column and centrifuged at 8,000 rpm for 1 min. The column was washed with wash solution and centrifuged at 14,000 rpm for 1 min. Finally, the column was applied to 20 ul of elution solution and centrifuged at 14,000 rpm for 2 min, and the flow-through was stored at –80 ℃ for storage. RNA precipitates were quantified and qualified with Bioanalyzer Pico 6000 Chip (Agilent Technologies, Santa Clara, CA). The 10 ng of total RNA was provided to the cDNA library generation using SMARTer smRNA-Seq kit (Illumina Inc., San Diego, CA), according to the user manual. The cDNA library was subjected to sequencing using Truseq SBS Kit and HiSeq 2500 System (Illumina Inc., San Diego, CA), according to user guide.
2020-09-15 | GSE157916 | GEO