AGO5 | Argonaute 5

Product no: AS10 671

AS10 671  |  Clonality: Polyclonal  |  Host: Rabbit  |  Reactivity: Arabidopsis thaliana

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  • Product Info
  • Immunogen:

    peptide of Arabidopsis thaliana AGO5 UniProt: Q9SJK3, TAIR:AT2G27880

    Host: Rabbit
    Clonality: Polyclonal
    Purity: Immunogen affinity purified serum in PBS pH 7.4.
    Format: Lyophilized
    Quantity: 50 g
    Reconstitution: For reconstitution add 50 µl of sterile water
    Storage: Store lyophilized/reconstituted at -20C; 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: Immunofluorescebce (IF), Immunoprecipitation (IP), Western blot (WB)
    Recommended dilution: 1 : 100 (IF), 5 ug per gram floral tissue (IP), 1 : 1500 (WB)
    Expected | apparent MW:

    111 | 111 kDa

  • Reactivity
  • Confirmed reactivity: Arabidopsis thaliana
    Predicted reactivity:

    Arabidopsis thaliana

    Not reactive in: Hordeum vulgare, Solanum lycopersicum, Zea mays
  • Application Examples
  • Application example

    western blot using anti-AGO5 antibodies on Arabidopsis thaliana
    Arabidopsis thaliana total protein extracted by TCA-acetone precipitation (check protocol tab for details) from floral buds of Col-0 (which should be more enriched in AGO5) was saturated in 8M urea were separated on 10% 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 a dilution of 1:250 overnight at 4C 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, Dr Pawel Baster, University of Cambridge, United Kingdom
  • Additional Information
  • Additional information (application): AGO expression may be tissue specific and using floral tissue is recommended where most of the AGOs are expressed the highest, Use of proteasome inhibitors as MG132 can help to stabilize AGO proteins during extraction procedure
  • Background
  • Background:

    AGO5 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). Probably involved in antiviral RNA silencing.

  • 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.
  • Protocols
  • TCA/Acetone protein precipitation method for plant proteins


    • Grind plant tissue in a liquid nitrogen. 
    • Continue grinding with 10% TCA solution in acetone (ice cold).
    • Precipitate overnight in -20C.
    • Spin at 4°C for 1min, 17k rpm > wash with ice cold acetone until you obtain a white pellet.
    • Dissolve the pellet in buffer of choice (for example 8M urea containing 5mM DTT, or denaturate in SDS protein loading buffer for 10 min. at 70°C)
    • Clarify supernatant
    • Measure protein concentration. 
    • Proceed with a western blot.

    Example of a western blot result obtained with this method, which allows high protein load per well, can be found below. 



    western blot using anti-AGO4 antibodies


    360 µg/well of Arabidopsis thaliana protein extracted by TCA-acetone precipitation from floral tissue 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 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 (DayLight 800) 1:5000 dilution for 30 min. at RT with agitation and washed 1X with TBSTT for 15 min, 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

    BACK to Plant Protocols page
  • Reviews:
  • Reviews
    Below, you can grade the product on a scale from 0 to 5.
    Please also provide information about species, application, dilution and obtained result for the reviewed antibody.
    Your name will be displayed as the sender.
    Number of reviews: (1)
    Guilherme Silva Martins | 2017-05-04
    0 | 0
    AGO5 protein is a really tricky protein that required good antibody for its detection. Agrisera AGO5-antibody demonstrate to be good for AGO5 detection. The reason why I gave this grade it is because this antibody detect a lot of non-target protein resulting into a lot of unespecific bands. Although the unespecific bands, we could detect our protein and Agrisera antibody showed to be the better one when we compare with others company.Thank you for this product

Accessories

AS09 602 |  Clonality: Polyclonal | Host: Goat | Reactivity: Rabbit IgG (H&L)

213 €
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AS16 ECL-S | extreme low femtogram detection

This product can be purchased in 3 different volumes:

AS16 ECL-S-10, 10 ml. Trial size limited to one per customer

AS16 ECL-S-100, 100 ml

AS16 ECL-S-1L, 1L

Choose the appropriate volume in the drop down menu to the right

From 17 €
AS18 4248 |  Clonality: Polyclonal  |  Host: Rabbit  |  Reactivity: Arabidopsis thaliana

This set contains:
- 4 Anti-AGO antibodies of your choice, chosen from a drop down menu below
- Matching secondary antibody, HRP conjugated, min. 1: 25 000, 1h/RT (AS09 602 - trial)
- Chemiluminescent reagent, femtogram detection range AgriseraECLSuperBright

1167 €
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