Team:Alberta/Notebook/ReusablePlates

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=Creating Inexpensive and Reusable Plates=
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===Project Timeline: Click on an image to see more information===
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<html>
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<img src="https://static.igem.org/mediawiki/2010/0/09/PlatesTimeline.jpg" border="0" usemap="#Plates">
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<map name="Plates">
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<area shape="rect" coords="0, 0, 100, 83" href="https://2010.igem.org/wiki/index.php?title=Team:Alberta/Notebook/ReusablePlates#Scintered_Plastic_Plates">
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<area shape="rect" coords="191, 0, 290, 82" href="https://2010.igem.org/wiki/index.php?title=Team:Alberta/Notebook/ReusablePlates#L.B.O.">
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<area shape="rect" coords="367, 25, 466, 72" href="https://2010.igem.org/wiki/index.php?title=Team:Alberta/Notebook/ReusablePlates#Jello_and_Gelatine_Plates">
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</map>
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</html>
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----------
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The equipment and supplies required to sterilize and make plates are expensive and not usually available to a high school lab.  We attempted to create plates which could easily be sterilized by a microwave or bleach.
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==Scintered Plastic Plates==
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Scintered Plastic Supplier: SPC technologies Ltd.
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We used 3 types of scintered plastic:
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:*3.0mm Ultra-Fine PE sheet(14um pore-size)
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::**ultra-fine grade should be a bacterial barrier
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:*6.0mm Fine-Grade PE Sheet(30um pore-size)
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::**fine grade should also be a bacterial barrier but it was too hard to cut
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:*4.5mm Medium Grade PE Sheet(88um pore-size)
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Procedure:
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#tested plastic's ability to absorb LB
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:*June 22, 2010 - added 10ml of LB and 40ul chloramphenicol to plates made from the 3.0mm and 4.5mm scintered plastics.  Incubated in a 37C warm room overnight.
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:*June 23, 2010 - 7ml LB left in 3.0mm ultra-fine and 8ml left in 4.5mm medium-grade
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#Streaking on Plastic
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:*June 23, 2010 - streaked both types of scintered plastic with RFP-containing colonies(after transferring plastic into new petri dishes).  Left in 37C warm room overnight.
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:*June 24, 2010 - no growth after incubation, plastic was very dry
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==L.B.O==
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===Creation of "L.B.O.": a deodorant stick of LB agar===
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*June 24, 2010 - All of the following steps were performed in a safety cabinet under sterile conditions.
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# disassembled a Degree deodorant stick and soaked in ethanol
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# removed the raising platform and covered it with Parafilm
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# coated the insides of the tube with mineral oil
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# removed platform and poured LB agar into the base of the stick, waited until it solidified
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# the Parafilmed platform was put back into the stick and lowered maximally
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# LB agar was poured on top of the platform, until the tube was full, waited until it solidifed
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# The top of the LB agar was sliced off with an ethanol-sterilized knife
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# 400ul of transformed competent cells were plated on the stick as if it was a standard LB agar plate
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# L.B.O. stick was incubated overnight at 37C
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<html>
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<center>
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<img src="http://farm5.static.flickr.com/4103/5116068476_a7c90747ba_m.jpg" width="240" height="180" alt="LBO" />
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</center>
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</html>
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Observations:
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:* although the knife cut through the LB agar easily, it was difficult to produce a completely flat surface
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:* solidified LB agar was effectively raised and lowered using the dial
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:* after incubation, a bacterial lawn was observed and a distinct E.coli smell was present
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*June 29, 2010
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# Followed procedure from June 26, 2010 to put new LB agar into the L.B.O. tube
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# Approximately 1cm was sliced off the top of the new LB agar with an ethanol-sterilized razor blade
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# L.B.O. stick was incubated overnight at 37C
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Observations:
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:* no growth observed, tube must have been sterile
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*June 30, 2010
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# Streaked green colony from a plate of Cambridge parts
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# L.B.O. stick was incubated overnight at 37C
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Observations:
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:* green streak and individual green colonies observed, along with a distinct E.coli smell
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*July 5, 2010
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# Approximately 1cm was sliced off the top of the used LB agar with an ethanol-sterilized razor blade
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# Streaked red, RFP-containing colony onto the LB agar surface.
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# L.B.O. stick was incubated overnight at 37C
 +
Observations:
 +
:* only a red streak and individual red colonies observed, along with a distinct E.coli smell
 +
:* no remnants of green colonies visible
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==Jello and Gelatine Plates==
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Used two types of Jello:
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#Jell-o brand, raspberry flavor:
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:* sweetened with aspartame (no sugar added)
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:*125mg sodium per 2.5g
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:*1g protein per 2.5g
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:2. Knox brand Gelatine
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Procedure (July 26, 2010):
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Jello:
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:* Stir two cups boiling water into 10g powder until dissolved
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:*pour into petri dishes
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:*chill for 2 hours until set
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Gelatine:
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:*stir 175ml milli-Q water into 15ml gelatine
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:*stir in 175ml hot milli-Q and pour into petri dishes
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:*put in cold room until set (approx 2hours)
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After 2 hours (Gelatine and Jello):
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:*Spread BBa_k274003(dark green stain of E.coli) onto 2 jello and 2 gelatine plates
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:*incubate overnight at 37C
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Results:
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Both Jello and Gelatine plates had liquefied  and no growth was seen.
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{{Team:Alberta/endMainContent}}

Latest revision as of 03:59, 28 October 2010

TEAM ALBERTA

Creating Inexpensive and Reusable Plates

Project Timeline: Click on an image to see more information


The equipment and supplies required to sterilize and make plates are expensive and not usually available to a high school lab. We attempted to create plates which could easily be sterilized by a microwave or bleach.

Scintered Plastic Plates

Scintered Plastic Supplier: SPC technologies Ltd.

We used 3 types of scintered plastic:

  • 3.0mm Ultra-Fine PE sheet(14um pore-size)
    • ultra-fine grade should be a bacterial barrier
  • 6.0mm Fine-Grade PE Sheet(30um pore-size)
    • fine grade should also be a bacterial barrier but it was too hard to cut
  • 4.5mm Medium Grade PE Sheet(88um pore-size)

Procedure:

  1. tested plastic's ability to absorb LB
  • June 22, 2010 - added 10ml of LB and 40ul chloramphenicol to plates made from the 3.0mm and 4.5mm scintered plastics. Incubated in a 37C warm room overnight.
  • June 23, 2010 - 7ml LB left in 3.0mm ultra-fine and 8ml left in 4.5mm medium-grade
  1. Streaking on Plastic
  • June 23, 2010 - streaked both types of scintered plastic with RFP-containing colonies(after transferring plastic into new petri dishes). Left in 37C warm room overnight.
  • June 24, 2010 - no growth after incubation, plastic was very dry

L.B.O

Creation of "L.B.O.": a deodorant stick of LB agar

  • June 24, 2010 - All of the following steps were performed in a safety cabinet under sterile conditions.
  1. disassembled a Degree deodorant stick and soaked in ethanol
  2. removed the raising platform and covered it with Parafilm
  3. coated the insides of the tube with mineral oil
  4. removed platform and poured LB agar into the base of the stick, waited until it solidified
  5. the Parafilmed platform was put back into the stick and lowered maximally
  6. LB agar was poured on top of the platform, until the tube was full, waited until it solidifed
  7. The top of the LB agar was sliced off with an ethanol-sterilized knife
  8. 400ul of transformed competent cells were plated on the stick as if it was a standard LB agar plate
  9. L.B.O. stick was incubated overnight at 37C

LBO
Observations:

  • although the knife cut through the LB agar easily, it was difficult to produce a completely flat surface
  • solidified LB agar was effectively raised and lowered using the dial
  • after incubation, a bacterial lawn was observed and a distinct E.coli smell was present
  • June 29, 2010
  1. Followed procedure from June 26, 2010 to put new LB agar into the L.B.O. tube
  2. Approximately 1cm was sliced off the top of the new LB agar with an ethanol-sterilized razor blade
  3. L.B.O. stick was incubated overnight at 37C

Observations:

  • no growth observed, tube must have been sterile
  • June 30, 2010
  1. Streaked green colony from a plate of Cambridge parts
  2. L.B.O. stick was incubated overnight at 37C

Observations:

  • green streak and individual green colonies observed, along with a distinct E.coli smell
  • July 5, 2010
  1. Approximately 1cm was sliced off the top of the used LB agar with an ethanol-sterilized razor blade
  2. Streaked red, RFP-containing colony onto the LB agar surface.
  3. L.B.O. stick was incubated overnight at 37C

Observations:

  • only a red streak and individual red colonies observed, along with a distinct E.coli smell
  • no remnants of green colonies visible

Jello and Gelatine Plates

Used two types of Jello:

  1. Jell-o brand, raspberry flavor:
  • sweetened with aspartame (no sugar added)
  • 125mg sodium per 2.5g
  • 1g protein per 2.5g
2. Knox brand Gelatine

Procedure (July 26, 2010): Jello:

  • Stir two cups boiling water into 10g powder until dissolved
  • pour into petri dishes
  • chill for 2 hours until set

Gelatine:

  • stir 175ml milli-Q water into 15ml gelatine
  • stir in 175ml hot milli-Q and pour into petri dishes
  • put in cold room until set (approx 2hours)

After 2 hours (Gelatine and Jello):

  • Spread BBa_k274003(dark green stain of E.coli) onto 2 jello and 2 gelatine plates
  • incubate overnight at 37C

Results: Both Jello and Gelatine plates had liquefied and no growth was seen.




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