2024-07-06
ENA first public
ERC000011
ENA-CHECKLIST
SAMEA114530576
External Id
Institute of Molecular Genetics of the Czech Academy of Sciences Laboratory of Genomics and Bioinformatics
INSDC center name
2023-10-24T14:35:27Z
INSDC last update
public
INSDC status
ERS16511912
SRA accession
E-MTAB-13483:Sirt1null-3
Submitter Id
10 to 12
week
age
ArrayExpress
broker name
not collected
collection date
house mouse
common name
Protocols: 10-12 weeks old female mice were used as oocyte donors. For the collection of in vivo ovulated (IVO) matured oocytes, PMSG administration was followed by hCG 48 h later; 15–16 h post-hCG injection, ovulated cumulus-oocyte complexes were flushed from the fallopian tube and treated with 0.1 mg/ml hyaluronidase (Sigma-Aldrich, #H3506) in M2 medium for 5 min to remove cumulus cells. Matured IVO oocytes were incubated in potassium simplex optimisation medium (KSOM), modified by 0.1 % of bovine serum albumin (BSA; mKSOM). For the preparation of activating medium, the mKSOM was supplemented with 2 mM EGTA, 10 mM SrCl2, and 5 μg/mL cytochalasin B, for 5.5 h at 37 °C and 5 % CO2. Cycloheximide (CHX; 15 µg/ml) was used for the inhibition of proteosynthesis during the oocyte activation. Afterwards, embryos were moved into pure mKSOM and cultured for 24 to cleaved embryos stages. For each sample, approximately 50 oocytes were collected in TRIzol reagent (Invitrogen, USA) and homogenised, using TissueLyser LT (Qiagen, The Netherlands) for 5 min at 50 Hz. Chloroform was used for phase separation; the aqueous phase was mixed 1:1 with 70 % ethanol and applied to RNeasy MinElute spin columns (Qiagen). Wash and elute steps were done according to the column manufacturer´s instructions. Isolated RNA was processed, using the Takara Smarter Stranded Total RNA-seq Kit v2 Pico Input Mammalian (PN:634413), according to the manufacturer's instructions.
description
adult
developmental stage
Zp3-Cre negative
genotype
not collected
geographic location (country and/or sea)
1
individual
not applicable
isolate
Mus musculus
organism
Mus musculus
organism
oocytes
organism part
Mus musculus
scientific_name
female
sex
C57BL/6-Tg(Zp3-cre)93Knw/J
strain
Sirt1null-3
title