Team:Chiba/Calendar-Home/21 October 2008

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20 October 2008 <|> 22 October 2008

Laboratory work

Team:Demo-Rs

Varying bacterial numbers

20 October~

    1. Pre-incubated Receiver(BBa_T9002(JW1908))was plated so as to produce about 1000 colonies.
  2. Sender(S03623) pre-incubation
    1. Sender:BBa_S03623(JW1908) was cultured in 50mL entrifuge tubes in 10mL of LB-Amp (37°C,12h)(2 tubes)
  3. Sender Wash
    1. Centrifuged 2 tubes containing(BBa_T9002(JW1908))at 20°C,3600rpm for 6min and discarded supernatant.
    2. Added 10mL LB-Amp to each tube.
    3. Repeated wash twice.
  4. Creating bacterial plates
    1. LB-Amp pre-cultured Sender(BBa_S03623(JW1908)) tube 1 (10mL) was mixed with LB-Amp-agar(50°C)(10ml)to produce sender containing bacterialplate-1.
    2. LB-Amp pre-cultured Sender(BBa_S03623(JW1908)) tube 2(100μl)was mixed with LB-Amp(9.9ml) and diluted 100-fold. 10ml of this solution was mixed with LB-Amp-agar(50°C)(10ml) and created Sender(BBa_S03623(JW1908))containing bacterial plate-2.
    3. LB-Amp pre-cultured Sender solution-2(10μl) and LB-Amp(9.99ml)was mixed to dilute 1000-fold.10ml of this solution and LB-Amp-agar(50°C)(10ml) was mixed to create Sender(BBa_S03623(JW1908) containing bacterial plate-3
  5. Lifted with nitrocellulose
    1. Receiver(BBa_T9002(JW1908))colony was transfered to a nitrocellulose filter and placed on each of Sender(BBa_S03623(JW1908))containing bacterial plate (1~3) and Sender-absent negative control plate (t=0). Determined the time required for the colonies to fluoresce depending on the bacterial concentration (100 and 1000-fold dilution).
  6. Method to detect fluorescence
    1. Plates cultured at 37°C were exposed to UV (312nm) light once every 30 minutes to observe GFP fluorescence.
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