Team:UNIPV-Pavia/Calendar/July/settimana2

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<html><p align="center"><font size="4"><b>JULY: WEEK 2</b></font></p></html><hr><br>
 
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<table class="menu" border="0" width="100%">
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[[Team:UNIPV-Pavia/Calendar/July/settimana1|Week 1]]
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</td>
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<td align="center" style="padding:0; height:20px">
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[[Team:UNIPV-Pavia/Calendar/July/settimana2|Week 2]]
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<td align="center">
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[[Team:UNIPV-Pavia/Calendar/July/settimana3|Week 3]]
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<td align="center">
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[[Team:UNIPV-Pavia/Calendar/July/settimana4|Week 4]]
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</td>
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<td align="center">
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[[Team:UNIPV-Pavia/Calendar/July/settimana5|Week 5]]
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</td>
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</table>
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<br>
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<html><p align="center"><font size="4"><b>JULY: WEEK 2</b></font></p></html><hr><br>
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<html><a name="indice"/></html>
==July, 5th==
==July, 5th==
 +
LB agar plates prepared:
 +
*LB+Amp 100 (500ml)
 +
*LB+Cm 34 (500ml)
 +
*LB (250ml)
 +
*LB+Cm 12.5 (250ml)
 +
 +
<div align="right"><small>[[#indice|^top]]</small></div>
==July, 6th==
==July, 6th==
 +
We decided to perform a TECAN test to evaluate the strength of some promoters belonging to the Anderson Promoters Collection. Inoculum from glycerol stocks (8ul in 5ml LB) for:
 +
*<partinfo>BBa_J23101</partinfo>
 +
*<partinfo>BBa_J23110</partinfo>
 +
*<partinfo>BBa_J23116</partinfo>
 +
*<partinfo>BBa_J23118</partinfo>
 +
*<partinfo>BBa_J23114</partinfo>
 +
*<partinfo>BBa_B0033</partinfo>
 +
 +
TECAN experiment started at 10:20am.
 +
 +
In the morning one single colony was picked for BW53474 E. coli strain and glycerol stock was prepared after about 7 hours (37°C 220rpm)
 +
 +
Inoculum of:
 +
*BW53474 (falcon containing culture for glycerol stock was re-filled with 5ml LB)
 +
*MG1655 (from glycerol stock)
 +
*BW25141 (from glycerol stock)
 +
*BW25142 (from glycerol stock)
 +
 +
<div align="right"><small>[[#indice|^top]]</small></div>
==July, 7th==
==July, 7th==
 +
Competent cells preparation for:
 +
*MG1655
 +
*BW53474
 +
*BW25141
 +
*BW25142
 +
 +
PCR was performed to amplify Phasins:
 +
 +
All parts were correct!! :)
 +
 +
<div align="right"><small>[[#indice|^top]]</small></div>
==July, 8th==
==July, 8th==
-
Transformation of 4ug of miniprepped <partinfo>BBa_J23118</partinfo> into home-made competent cells
+
Transformation of 1ul (~4ng) of miniprepped <partinfo>BBa_J23118</partinfo> into 100ul of home-made competent cells
*MG1655
*MG1655
*BW25141
*BW25141
*BW25142
*BW25142
*BW53474
*BW53474
-
*DH5alpha (as control)
+
*DH5-alpha (as control)
to verify the transformation efficiency.
to verify the transformation efficiency.
-
Transformed bacteria are plated on LB+Amp plates and left ON in oven, 37°C.
+
Transformed bacteria were plated on LB+Amp and left overnight in oven, 37°C.
 +
 
 +
<div align="right"><small>[[#indice|^top]]</small></div>
==July, 9th==
==July, 9th==
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<td width="20%" valign="top">
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We checked the presence of colonies in plates.
 +
All plates showed red colonies!
 +
 
 +
{|align="center"
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| [[Image:MG_unipv10_efficiency.jpg|200px|thumb|center|MG1655 plate (with satellite colonies)]] || [[Image:BW25141_unipv10_efficiency.jpg|200px|thumb|center|BW25141 plate]] || [[Image:BW25142_unipv10_efficiency.jpg|200px|thumb|center|BW25142 plate]]
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|-
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|[[Image:BW53474_unipv10_efficiency.jpg|200px|thumb|center|BW53474 plate]] || [[Image:DH5-alpha_unipv10_efficiency.jpg|200px|thumb|center|DH5-alpha plate]]
 +
|-
 +
|}
 +
 
 +
We calculated (thanks Nicolò) efficiency as #colonies/ug DNA
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{|border="1" align="center"
 +
!Strain || #colonies || Efficiency
 +
|-
 +
| MG1655 || 379 || ~10^5
 +
|-
 +
| BW25141 || 1431 || ~10^5
 +
|-
 +
| BW25142 || 2374 || ~10^5
 +
|-
 +
| BW53474 || 9984 || ~10^6<br/>(very difficult to count correctly)
 +
|-
 +
| DH5alpha ||  7475<br/>(only 100ul of cells plated)  || ~ 10^6<br/>(very difficult to count correctly,<br/>previous tests showed an efficiency of ~10^8)
 +
|}
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<div align="right"><small>[[#indice|^top]]</small></div>
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<table border="0" width="100%" height="100%">
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<!-- table previous next week -->
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<tr align="right"><td height="15">
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<br><br>
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[[Team:UNIPV-Pavia/Calendar|Calendar]]
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<table border="0" width="100%" class="menu">
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</td></tr>
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<tr>
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<tr align="right"><td height="15">
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<td align="left">[[Team:UNIPV-Pavia/Calendar/July/settimana1| Previous week]]</td>
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[[Team:UNIPV-Pavia/Calendar/July|July]]
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<td align="right">[[Team:UNIPV-Pavia/Calendar/July/settimana3| Next week]]</td>
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</td></tr>
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</tr>
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<tr align="right"><td height="15">
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[[Team:UNIPV-Pavia/Calendar/July/settimana1|week 1]]
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</td></tr>
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<tr align="right"><td height="15">
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[[Team:UNIPV-Pavia/Calendar/July/settimana2|week 2]]
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</td></tr>
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<tr align="right"><td height="15">
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[[Team:UNIPV-Pavia/Calendar/July/settimana3|week 3]]
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</td></tr>
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<tr align="right"><td height="15">
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[[Team:UNIPV-Pavia/Calendar/July/settimana4|week 4]]
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</td></tr>
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</table>
</table>
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<!-- fine table previous next week -->
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</td>
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<td width="15%" align="right" valign="top">
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{{UNIPV-Pavia/menu_mesi}}
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Latest revision as of 16:53, 24 October 2010
Home
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Week 1

Week 2

Week 3

Week 4

Week 5


JULY: WEEK 2


July, 5th

LB agar plates prepared:

  • LB+Amp 100 (500ml)
  • LB+Cm 34 (500ml)
  • LB (250ml)
  • LB+Cm 12.5 (250ml)

July, 6th

We decided to perform a TECAN test to evaluate the strength of some promoters belonging to the Anderson Promoters Collection. Inoculum from glycerol stocks (8ul in 5ml LB) for:

  • <partinfo>BBa_J23101</partinfo>
  • <partinfo>BBa_J23110</partinfo>
  • <partinfo>BBa_J23116</partinfo>
  • <partinfo>BBa_J23118</partinfo>
  • <partinfo>BBa_J23114</partinfo>
  • <partinfo>BBa_B0033</partinfo>

TECAN experiment started at 10:20am.

In the morning one single colony was picked for BW53474 E. coli strain and glycerol stock was prepared after about 7 hours (37°C 220rpm)

Inoculum of:

  • BW53474 (falcon containing culture for glycerol stock was re-filled with 5ml LB)
  • MG1655 (from glycerol stock)
  • BW25141 (from glycerol stock)
  • BW25142 (from glycerol stock)

July, 7th

Competent cells preparation for:

  • MG1655
  • BW53474
  • BW25141
  • BW25142

PCR was performed to amplify Phasins:

All parts were correct!! :)

July, 8th

Transformation of 1ul (~4ng) of miniprepped <partinfo>BBa_J23118</partinfo> into 100ul of home-made competent cells

  • MG1655
  • BW25141
  • BW25142
  • BW53474
  • DH5-alpha (as control)

to verify the transformation efficiency.

Transformed bacteria were plated on LB+Amp and left overnight in oven, 37°C.

July, 9th

We checked the presence of colonies in plates. All plates showed red colonies!

MG1655 plate (with satellite colonies)
BW25141 plate
BW25142 plate
BW53474 plate
DH5-alpha plate

We calculated (thanks Nicolò) efficiency as #colonies/ug DNA

Strain #colonies Efficiency
MG1655 379 ~10^5
BW25141 1431 ~10^5
BW25142 2374 ~10^5
BW53474 9984 ~10^6
(very difficult to count correctly)
DH5alpha 7475
(only 100ul of cells plated) 		~ 10^6
(very difficult to count correctly,
previous tests showed an efficiency of ~10^8)



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