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Sample GSM1602585 Query DataSets for GSM1602585
Status Public on Feb 06, 2015
Title E14.5-RA-3
Sample type SRA
 
Source name heart - right atrium
Organism Mus musculus
Characteristics strain background: mixed
developmental stage: Embryonic Day 14.5
genotype/variation: Hcn4-GFP
tissue: heart; right atrium
Treatment protocol Embryos or hearts were removed intact, washed with cold PBS, and immediately embedded in OCT and stored at -20 °C until sectioning. Tissue was sectioned at a thickness of 8 microns onto membrane-coated slides (MembraneSlide NF 1.0 PEN, Zeiss Microscopy, Gottingen, Germany). For laser capture, slides were thawed to room temperature until evaporation of moisture (approximately 1 minute), placed on the microscope stage of a PALM Micro-Beam inverted microscope with LCM capability (Zeiss). The sinus node tissue and right atrial tissue were identified visually and outlined manually with the microscope user interface (Online Video 2). Laser power and catapult energy were optimized prior to the experiment and varied from experiment-to-experiment. After each experiment, sections were stained with DAPI and anti-Hcn4, mounted, and visualized to confirm accurate dissection of the region of interest.
Growth protocol Genetically modified mice were maintained on a mixed background.
Extracted molecule total RNA
Extraction protocol Total RNA was prepared from regions of interest isolated with laser capture using the PicoPure Kit (Arcturus) according to manufacturers instructions.
RNA-seq libraries were prepared with the Ovation RNA-Seq System V2 kit (NuGEN) according to manufacturers instructions.  Briefly, total RNA is reverse-transcribed using a combination of random hexamers and a poly-T chimeric primer, followed by second strand synthesis using DNA polymerase. The cDNA was then amplified using single primer isothermal amplification (SPIA). Libraries from the SPIA amplified cDNA were made using the Ultralow DR library kit  (NuGEN) according to manufacturers instructions.  The RNA-seq libraries were analyzed by Bioanalyzer (Agilent) and quantified by QPCR (KAPA).
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 2000
 
Description RA.E14.5.2
Data processing Short read assembly was performed using TopHat v2.0.6 against mm8 build of the genome.
Gene counts were determined using Rsamtools and sample normalization was performed using edgeR.
Differential expression was performed using edgeR.
Genome_build: mm8
Supplementary_files_format_and_content: text-tab delimited with rows as genes and columns as samples; contains normalized counts as output by edgeR
 
Submission date Feb 05, 2015
Last update date May 15, 2019
Contact name Vasanth Vedantham
E-mail(s) vedanthamv@medicine.ucsf.edu
Organization name University of California, San Francisco
Department Medicine-Cardiology
Street address 505 Parnassus Avenue, M1176D
City San Francisco
State/province CA
ZIP/Postal code 94143
Country USA
 
Platform ID GPL13112
Series (1)
GSE65658 Mouse sinoatrial node transcriptome
Relations
SRA SRX864570
BioSample SAMN03328585

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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