AGO6 | Argonaute 6

AS10 672 | Clonality: Polyclonal | Host: Rabbit | Reactivity: Arabidopsis thaliana

Product Information

Immunogen

KLH-conjugated peptide derived from Arabidopsis thaliana AGO6 UniProt: O48771 TAIR:At2g32940

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 Western blot (WB)

Recommended dilution 1 : 1000 (WB)

Expected | apparent MW 116,4 | 99 kDa

Reactivity

Confirmed reactivity Arabidopsis thaliana

Predicted reactivity

Arabidopsis thaliana

Not reactive in Zea mays

Application examples

Application examples Application example

western blot using anti-AGO6 antibodies'

360 µg/well of Arabidopsis thaliana protein extracted from leaf using TCA-protein precipitation (for method check protocol tab) and saturated in 8M urea were separated on 15% SDS-PAGE and blotted for 1hour to 0.2 µm nitrocellulose at 100V using wet transfer system. Blots were blocked with 0.5% cold fish gelatin for 1hr at room temp with agitation. Blot was incubated in the primary antibody (anti-H3) at a dilution of 1:2500 for an hour at RT with agitation. The blots were washed with 3X 15min TBS-TT at RT with agitation. Blots as incubated in the secondary antibody (goat anti-rabbit DyLight® 800 conjugated, AS12 2460, Agrisera) 1:5000 dilution for 30min at RT with agitation and washed 1X with TBSTT for 15min, 1X with TBST for 15min before scanning with the ODyssey IRD scanner.

Courtesy of Dr. Betty Chung and Pawel Baster, University of Cambridge, United Kingdom

Additional information

There can be a reaction with NEB broad range prestained protein ladder depending upon secondary antibody used.

AGO6 accumulates to lower levels in the nrpd1a mutant (Havecker at el. 2010). In addition, a non-specific band migrates to a similar location and thus protein was run on a 6% denaturing gel to obtain good separation.

If other secondary antibody is used than in the figure below, cross-reacting pattern can be different.

AGO6 expression is cell specific, Seedlings can be used as a negative control

Background

AGO6 (Argonaute 6) belongs to a group of argonaute proteins which are catalytic component of the RNA-incudes silencing complex (RISC). This protein complex is responsible for the gene silencing (RNAi).

Product citations

Selected references Oliver & Martinez. (2021) Accumulation dynamics of ARGONAUTE proteins during meiosis in Arabidopsis. Plant Reprod. 2021 Nov 23. doi: 10.1007/s00497-021-00434-z. Epub ahead of print. PMID: 34812935.
Sprunck et al. (2019). Elucidating small RNA pathways in Arabidopsis thaliana egg cells. http://dx.doi.org/10.1101/525956
Havecker el al. (2010). The RNA-directed DNA methylation Arabidopsis Argonautes functionally diverge based on expression and interaction with target loci. Plant Cell. 2010 Feb;22(2):321-34. doi: 10.1105/tpc.109.072199.