Data · dataset · 2022
Telomeric noncoding RNA promotes mouse embryonic stem cell self-renewal through inhibition of TCF3 activity
Listed in ScienceDB
Immunofluorescence staining.
Description
The cells were fixed in 4% parafor-maldehyde for 20 min and incubated at 37°C in blocking buffer (PBS containing 5% BSA and 0.2% Triton X-100). Then, the cells wereincubated in the presence of primary antibodies at 4°C overnight and were washed three times in PBS.
Finally, the cells were incubated with an Alexa Fluor 488 (1:1,000, catalog no. A-11034, Invitrogen) or Alexa Fluor 594 (1:1,000, catalog no. A-11032; Invitrogen) secondary antibody for 1 h at 37°C. Nuclei were stained with Hoechst 33342 (1:10,000, catalog no. H1399; Invitrogen). The primary antibodies were against Nanog (1:500, catalog no. 14295-1-AP; Proteintech), Oct4 (1:200, catalog no. SC-5279; Santa Cruz), and SSEA1 (1:100, catalog no. SC-21702; Santa Cruz).Western blotting. mESCs were lysed in ice-cold RIPA cell buffer (Beyotime Biotechnology, China) supplemented with protease inhibitors (Transgene Biotech).
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Total protein was separated by a 10% PAGE gel (Bio-Rad) and was electrotransferred onto a polyvinylidene difluoride membrane. The primary antibodies used for probing were TCF3 (1:200, catalog no. SC-8635; Santa Cruz), FLAG (1:1,000, catalog no. F1804; Sigma), and α-tubulin (1:1,000, catalog no. SC-8035; Santa Cruz).The following secondary antibodies were used:goat anti-mouse IgG(H +L), horseradish peroxidase (HRP) conjugate(catalog no. HS201-01; Transgene Biotech) and donkey anti-goat IgG(H+L), HRP conjugate (catalog no. SC-2020; Santa Cruz).
The signals were detected using Pro-Light HRP Kit (PA112-02, Tiangen Biotech, China). The density of the protein bands was quantified by ImageJ (NIH, Bethesda, MD).Accession number. Data and details of transcriptome-resequencing are available in the Gene Expression Omnibus under accession number GSE103933.Statistical analysis.
All data are reported as means±SD. Student’s t-test was used to determine the significance of differences in comparisons. Values of P≤0.05 were considered as statistically significant.
Links
Where it is published
- DOI doi.org/10.57760/sciencedb.03358 ↗
DOI / persistent id · from scidb cn
Catalogue records · 1
- OAI-PMH record scidb.cn/oai?verb=GetRecord&metadataPrefix=oai_dc&identifier=10.57760%2… ↗
metadata API · from scidb cn
Topics
- From keywords
- Earth & Environmental Science · Engineering · Humanities · Life Sciences · Social Science
Provenance · 1 source records, 10 field assertions
| Source | Key | Last seen | Raw |
|---|---|---|---|
| ScienceDB | 10.57760/sciencedb.03358 | 8 d ago | JSON v1 |
| Field | Assertion | Extractor | Evidence |
|---|---|---|---|
| access_level | source · scidb cn | connector:scidb_cn@1.0.0 | |
| concepts[field].local:field:earth-environmental | mapping · scidb cn | connector:scidb_cn@1.0.0 | |
| concepts[field].local:field:engineering | mapping · scidb cn | connector:scidb_cn@1.0.0 | |
| concepts[field].local:field:humanities | mapping · scidb cn | connector:scidb_cn@1.0.0 | |
| concepts[field].local:field:life-sciences | mapping · scidb cn | connector:scidb_cn@1.0.0 | |
| concepts[field].local:field:social-science | mapping · scidb cn | connector:scidb_cn@1.0.0 | |
| description | source · scidb cn | connector:scidb_cn@1.0.0 | /metadata/dc/description |
| license | source · scidb cn | connector:scidb_cn@1.0.0 | /metadata/dc/rights |
| publication_date | source · scidb cn | connector:scidb_cn@1.0.0 | |
| title | source · scidb cn | connector:scidb_cn@1.0.0 | /metadata/dc/title |