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Sample GSM1373903

Query DataSets for GSM1373903

Status

Public on Nov 22, 2021

Title

no RNA negative control_1

Sample type

SRA

Source name

no RNA

Organism

synthetic construct

Characteristics

control type: negative

Growth protocol

48h post injection of 5 IU pregnant mare serum fully grown oocytes at the germinal vesicle stage were obtained from ovaries of 6-8 weeks old F1(C57BL/6×DBA) female mice and transferred to M2 medium without BSA, supplemented with 0.2 mM 3-isobutyl-1-methylxanthine (IBMX; Sigma-Aldrich). After 10 min of Hoechst 33342 (0.05 μg/ml) incubation SN type GVOs were injected with morpholinos designed against Tet1-3 enzymes (Gene Tools, 100 μM, see table S1) co-injected with Dextran-tetramethyl-rhodamine (Invitrogen, 3000 MW, 100 μg/ml). Morpholino injected SN GVOs were washed in IMBX free α-MEM medium supplemented with 5% FBS and 10 ng/ml EGF and incubated in a humidified atmosphere with 6% CO2, 5% O2 and 89% N2 at 37 °C for 16-18 h to complete meiotic maturation. Sperm was isolated from the cauda epididymis of adult F1(C57BL/6×DBA) male mice and capacitated by pre-incubation for 1.5 h in pre-gassed HTF medium. In vitro matured oocytes were collected 16-18 h post injection of morpholinos. Mature MII oocytes were placed into a 100 μl drop of KSOM medium with capacitated sperm and incubated at 37 °C in a humidified atmosphere of 6% CO2, 5% O2 and 89% N2. 2-cell embryos were collected 29 hours following in vitro fertilization.

Extracted molecule

total RNA

Extraction protocol

2-cell embryos were collected 29 hours following in vitro fertilization and zona pellucida was removed from 2-cell embryos. Libraries were constructed using Clontech SMARTer Ultra Low Input RNA Kit, followed by NEBNext DNA Library Prep Master Mix Set

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2000

Description

neg1

Data processing

Reads were mapped to mouse reference genome mm10, depleted for sex chromosomes with STAR2.5.1b using default settings. Genes from RefSeq database that have at least ten reads in 50% of the embryos of one condition were used for further analysis. Differentially expressed genes were obtained using DEseq2.
Genome_build: mm10
Supplementary_files_format_and_content: FPM.txt: Tab-delimited text file includes FPM values.

Submission date

Apr 24, 2014

Last update date

Nov 22, 2021

Contact name

Julia Arand

E-mail(s)

[email protected]

Organization name

Medical University of Vienna

Street address

Schwarzspanierstr. 16