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  • Bruno Vellutini 16:17 on 2012/06/29 Permalink
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    L. ruber collection trip 

    Went to Aina’s beach to collect Lineus ruber. We brought ~100 L. ruber + white/pinkeyes and the long/thin species.

     
  • Bruno Vellutini 18:00 on 2012/06/20 Permalink
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    Sars Seminar 2012 

    [file]

    Question from audience: how knowing the developmental mechanisms of a larval structure and its fate in the adult tissues will help to understand the evolution of metazoan life cycles?

     
  • Bruno Vellutini 18:01 on 2012/06/06 Permalink
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    FITC probe and washes for fluorescent in situ 

    Chema developed Tt vasa using FITC probe for regular and fluorescent in situ and also Tt nanos. He washed the fluorescent ones according to the protocol from (10.1186/2041-9139-2-6) (basically the 20 minutes wash with detergent), which consists of:

    Fixed larvae hybridized with Digoxigenin-11-UTP (Roche Applied Science, Indiana- polis, IN, USA) labelled Tt-c-opsin and Fluorescein-12-UTP (Roche Applied Science, Indianapolis, IN, USA) labelled Tt-Pax6 probes at 2.5 ng/μl each, for 48 hours at 60°C. Larvae were incubated overnight with Anti-digoxigenin-POD antibody (Roche Applied Science, Indianapolis, IN, USA) at a 1:1000 dilution in 1× Blocking Reagent (Roche Applied Science, Indianapolis, IN, USA) overnight at 4°C, and stained with TSA Plus Cyanine 5 at a 1:100 dilution for 60 minutes at room temperature. Following staining, peroxidase activity was extinguished by incubation in 2.7% hydrogen peroxide in 1× PBS buffer for 90 minutes. Subsequently, larvae were incubated overnight with Anti-fluorescein-POD antibody (Roche Applied Science, Indianapolis, IN, USA) at a 1:1000 dilution in 1× Blocking Reagent (Roche Applied Science, Indianapolis, IN, USA) overnight at 4°C, and stained with TSA Plus Tetramethylrhodamine at a 1:100 dilution for 60 minutes at room temperature. Stained larvae were washed for 48 hours with multiple exchanges of 1× PBS buffer, and cleared with 80% glycerol.

    The washes apparently reduced the background, though also the signal. I’ll check the samples in the confocal.

    Bibliography

     
  • Bruno Vellutini 12:00 on 2012/06/06 Permalink
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    S9 Project Discussion 

    Presented my project discussion. Summary of the feedback:

    • List names next to the cited hypothesis.
    • Study better the larval theories and cnidaria.
    • Set aside implies non-evolutionary thinking, use it between “”.
    • Get better the fate of spiralian cells.
    • Compare expression patterns with other animals (vasa, for example).
    • Include citation for figures.
    • Modify figures to be clearer.
    • Polarize discussion of “set aside” cells and larval evolution.

    The source of the talk is here. First time using an HTML5/CSS3 based presentation. It is in Markdown and was converted to HTML Slidy using Pandoc. Resulting presentation was acceptable, but since the screen resolution of the monitor was different it messed the layout of some slides. Changing to another.

     
  • Bruno Vellutini 09:59 on 2012/06/01 Permalink
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    Got a cod sample and fixed it in 4% PFA at 4 °C overnight.

    02/06

    Stained the smaller samples with Phallacidin for 2h and prepared the slides for the confocal. It was harder then the jellyfish because the fibers are packed in bundles and do not spread flat in the slide. Tried disrupting them in various way and found out that the staining was not so good in the inner fibers. Took enough pictures anyway.

     
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