NASC Stock Detail Page
NASC ID: N2105608
Name: P7Y, mCITRINE-1xPH(OSBP)
ABRC stock number: CS2105608
Description: Each PIPline construct was transformed into C58 GV3101 Agrobacterium strain and selected on YEB media (5g/L beef extract; 1g/L yeast extract; 5g/L peptone; 5g/L sucrose; 15g/L bactoagar; pH 7.2) supplemented with antibiotics (Spectynomicin, Gentamycin). After two days of growth at 28°C, bacteria were collected using a single-use cell scraper, re-suspended in about 200mL of transformation buffer (10mM MgCl2; 5% sucrose; 0.25% silweet) and plants were transformed by dipping. Primary transformants (T1) were selected in vitro on the appropriate antibiotic/herbicide (glufosinate for CITRINE-tagged PIPline, hygromycin for CHERRY-tagged PIPline and kanamycin for CyPET-tagged PIPline). 24 independent T1s were selected for each PIPline. In the T2 generation at least 3 independent transgenic lines were selected using the following criteria when possible: i) good expression level in the root for detection by confocal microscopy, ii) uniform expression pattern, iii) single insertion line (1 sensitive to 3 resistant segregation ratio) and, iv) line with no obvious abnormal developmental phenotypes. PIPlines for which we could not find fluorescence out of 24 independent lines or that did not associated with any membrane compartments (including the PM) were not further analysed. The remaining PIPlines were rescreened in T3 using similar criteria as in T2 with the exception that we selected homozygous lines (100% resistant). At this step, we selected one transgenic line for each PIPline that was used for further analysis and crosses and that was distributed to the Arabidopsis stock centres.
Donation Date: 2014-10-06
| Donated by: | Ecole Normale Supérieure de Lyon Yvon Jaillais |
Donor Number: P7Y
Stock type: individual line
Material type: seed
Comment: Lipid-binding domains (LBD's): OSBP (Homo sapiens).
| Status | Price (£) |
|---|---|
| Available | £11.00 |
Germplasm Info
Mutagen: ema
Background: Col-0
Segregation status: homozygous
Growth requirement: none
Phenotype
geneticaly encoded biosensor for PI4P (phosphatidylinositol 4-phosphate) fused with mCITRINE (YFP), localized at the plasma membrane
References
Simon, M.L. et al. 2014. A multi-colour/multi-affinity marker set to visualize phosphoinositide dynamics in Arabidopsis. The Plant Journal 77(2):322-37.PubMed ID: 24147788.