MIP1 | Aquaporin, glycerol transport activity

AS15 2826 | Clonality: Polyclonal | Host: Rabbit | Reactivity: Chlamydomonas reinhardtii

MIP1 | Aquaporin, glycerol transport activity

Product Information

Immunogen KLH-conjugated synthetic peptide, derived from Chlamydomonas reinhardtii MIP1, UniProt: Q5VLJ9

Host Rabbit

Clonality Polyclonal

Purity Immunogen affinity purified serum in PBS pH 7.4.

Format Lyophilized

Quantity 50 µg

Reconstitution For reconstitution add 50 µl of sterile water

Storage Store lyophilized/reconstituted at -20°C; once reconstituted make aliquots to avoid repeated freeze-thaw cycles. Please remember to spin the tubes briefly prior to opening them to avoid any losses that might occur from material adhering to the cap or sides of the tube.

Tested applications Immunogold (IG), Western blot (WB)

Recommended dilution 1 : 200 (IG), 1 : 10 000 (WB)

Expected | apparent MW 31,5 | 43 kDa

Reactivity

Confirmed reactivity Chlamydomonas reinhardtii (strains CC3395, and UVM4 from Neupert et al., 2009. (Generation of Chlamydomonas Strains that Efficiently Express Nuclear Transgenes.)

Predicted reactivity Chlamydomonas reinhardtii

Not reactive in No confirmed exceptions from predicted reactivity are currently known

Application examples

Application examples application example

10 µg of total protein from 15ml cell suspension of Chlamydomonas reinhardtii CC3395, with a cell density of (~ 10 times 8) extracted with SDS buffer supplememnted with Protease inhibitor (cOmplete Ultra tablets, EDTA free, Roche, Mannheim), after removal of the cell debris via centrifugation, DTT was added (100mM final concentration) and were separated on 12 % SDS-PAGE and blotted 1h to PVDF using tank transfer. Blots were blocked with RotiBlock-Solution (Roth, Darmstadt) for 1h at room temperature (RT) or at 4°C ON with agitation. Blot was incubated in the primary antibody at a dilution of 1: 10,000 in TBS with 2% milk powder for 1h at RT with agitation. The antibody solution was decanted and the blot was washed 3 times for 10 minutes each with TBS-T at RT with agitation. Blot was incubated in secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated, from Sigma, St.Luis, MO) diluted to 1:80 000 in TBS tith 5% milk powder for 1h at RT with agitation. The blot was washed as above and developed for 1-2 min with ECL according to the manufacturer's instructions. Exposure time was 40 seconds.

Courtesy of Dr. Karin Komsic-Buchmann and Prof. Dr. Burkhard Becker, University of Cologne, Germany

Reactant: Chlamydomonas reinhardtii (Green Alga)

Application: Western Blotting

Pudmed ID: 33117171

Journal: Front Pharmacol

Figure Number: 1C

Published Date: 2020-10-30

First Author: Yoshida, M., Yamamiya, R., et al.

Impact Factor: 5.331

Open Publication

Construction of a C. reinhardtii transformant expressing hTRPA1. (A) PCR for actin and hTRPA1 using the genomic DNA from wild-type (WT) and TA1 cells as templates. The sizes expected from the genome database (https://phytozome-next.jgi.doe.gov) are 611 bp for actin and 490 bp for hTRPA1. (B) RT-PCR to confirm transcription. The expected sizes are 227 bp for actin and 490 bp for hTRPA1. (C) Western blot using anti-hTRPA1 antibody and anti-MIP1 (aquaporin) antisera. Presumed bands for hTRPA1 and MIP1 are indicated by the filled and open dots, respectively. Molecular masses are in kDa.

Additional information

As MIP1 is a membrane protein please use a high redox potential in your lysis buffer

Background

Background MIP1 (Aquaporin, glycerol transport activity) is localized to the contractile vacuole, which in most freshwater flagellates is used to expel excess water.

Product citations

Selected references Komsic-Buchmann et al. (2014). The Contractile Vacuole as a Key Regulator of Cellular Water Flow in Chlamydomonas reinhardtii. Eukaryotic cell (13), Issue 11: 1421-30.