Team:Paris Saclay/Notebook/August/7

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(→Thursday 7th August)
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===D - Lemon Sent===
===D - Lemon Sent===
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 +
==== PCR of pCola plasmide (Geranyl Synthase)====
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'by melanie'
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 +
we use the same protocol than yesterday but we have done 8 tubes to apply a gradient in the thermocycleur during the third step:
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#57°
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#56.3°
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#55.1°
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#53.3°
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#51°
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#49.3°
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#47.9°
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#47°
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 +
PCR protocol:
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98° --> 2min
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 +
5 PCR cycle:
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 +
{| class="wikitable centre" width="50%"
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|+
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|-
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! scope=col | time
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! scope=col | 10sec
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! scope=col | 20sec
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! scope=col | 45sec
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|-
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!temperature
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|98°
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|(depending on the tube - see the gradient)
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|72°
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|}
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 +
25 PCR cycle
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 +
{| class="wikitable centre" width="50%"
 +
|+
 +
|-
 +
! scope=col | time
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! scope=col | 10sec
 +
! scope=col | 20sec
 +
! scope=col | 45sec
 +
|-
 +
!temperature
 +
|98°
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|72°
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|72°
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|}
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 +
and last step: 72° during 10min
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 +
but we don't have any results
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 +
==== Test the odor of e.coli with pJBEI6409 plasmide====
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'by melanie'
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Transformation of competent cells 5mg1655) by electroporation (pJBEI6409)
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[https://2014.igem.org/Team:Paris_Saclay/Protocols/Transformation_of_competent_cells_by_electroporation]
=== Extraction of the Genomic DNA===
=== Extraction of the Genomic DNA===

Revision as of 15:33, 7 August 2014

Contents

Thursday 7th August

Lab work

C - Salicylate Inducible Suppressing System

DNA extraction from gel agarose

In progress


D - Lemon Sent

PCR of pCola plasmide (Geranyl Synthase)

'by melanie'

we use the same protocol than yesterday but we have done 8 tubes to apply a gradient in the thermocycleur during the third step:

  1. 57°
  2. 56.3°
  3. 55.1°
  4. 53.3°
  5. 51°
  6. 49.3°
  7. 47.9°
  8. 47°

PCR protocol: 98° --> 2min

5 PCR cycle:

time 10sec 20sec 45sec
temperature 98° (depending on the tube - see the gradient) 72°

25 PCR cycle

time 10sec 20sec 45sec
temperature 98° 72° 72°

and last step: 72° during 10min

but we don't have any results

Test the odor of e.coli with pJBEI6409 plasmide

'by melanie' Transformation of competent cells 5mg1655) by electroporation (pJBEI6409) [1]

Extraction of the Genomic DNA

by Juliette & Terry

From liquid culture made the EDIT !!!!

Protocol


Polymerase chain reaction

by Sean & Pierre


for this PCR, five tubes of each of the following Bio-bricks were prepared.

BBa_K517003

component volume
H2O 35.5μl
Phusion buffer 5X 10μl
dNTPs 2μl
iPS68bis 1μl
iPS69 1μl
DNA 1μl
Phusion enzyme 0.5μl


BBa_K762100


component volume
H2O 35.5μl
Phusion buffer 5X 10μl
dNTPs 2μl
iPS66 1μl
iPS67 1μl
DNA 1μl
Phusion enzyme 0.5μl

Tubes were placed in PCR machine with the following parameters.

Cycle step Temperature Time Cycle

Initial denaturation

98°C

1 min

1

Denaturation 98°C 15 s 25 - 30
Annealing 52°C 25 s 25 - 30
Extension 72°C 45 s 25-30
Final extension 72°C 10 min 1
Final extension 8°C hold 1


Members present:

  • Instructors and advisors: Alice.
  • Students: Eugene, Fabio, Hoang Vu, Juliette, Melanie, Pierre, Romain, Sean and Terry.

Back to the calendar

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