Team:University College London/LabBook/Week6

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== 6.2 ==
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== Wednesday (18.7.12)  ==
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'''Aim - Testing the transformation protocols:''' In light of the problems with cell competency, we decided to test whether the choice of protocol was contributing to the poor transformation results – especially as it would require another week to set up another cell line. We used a known competent cell line, and tested its competency against both of our cell lines, for Transformation Protocol 1 vs Transformation Protocol 2. Each sample was plated on an Ampicillin positive Agar and incubated overnight.
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(LOGO) Transformation - Protocol 1
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(LOGO) Transformation - Protocol 2
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'''Step ?:''' The table below describes the plates necessary for this investigation, each of which shoud carry Ampicillin antibiotic at a concentration of 50ug/ml.
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{| class="wikitable"
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|-
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! colspan="2" |Plates
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| rowspan="3" | Protocol 1 ||  W3100 – Original
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|  W3100 – Reinvigorated
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|  W3100 – Known Competent
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| rowspan="3" | Protocol 2 ||  W3100 – Original
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|  W3100 – Reinvigorated
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|  W3100 – Known Competent
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|}
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== Thursday 19.7.12 ==
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'''Aim  - Results from Transformation of pTop'''
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'''Results:''' Table below indicates there was significant growth from the Original W3100 cells, but not from other cell lines.
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== 6.3 ==
== 6.3 ==
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Revision as of 11:01, 1 August 2012

Contents

Tuesday 17.7.12

Aim -Testing the competency of the Reinvigorated Cell Line (W3110): After our original attempt to generate competency of the W3100 cell line was inadequate, we undertook a reinvigoration of the W3100 cell line (Expt 5.2). Here we aim to test the competency of the reinvigorated cell line using the same protocol as used in Expt 5.3, by evaluating their growth after transformation with a plasmid of a known high concentration (297ng/ul).

Method

(LOGO) Transformation Protocol 2

Step 1 – Thawing Cells: Use the reinvigorated W3100 cell line created in Week 5 (Expt 5.2)

Step 3 – Addition of BioBrick: To one 2ml eppendorf, add 1ul of pTop plasmid, and to another add nothing – this will be a control.

Step 7 – Adding Broth: SOC media was used, as it is preferred for this protocol.

Step 8 - Incubation: The table below indicates the Ampicillin concentration of the Agar gels.

Samples Volume Innoculated Antibiotic in Gel (conc)
Plasmid pTOP 36ul Ampicillin (50ug/ml)
Control Positive (No Plasmid) 36ul No Antibiotic
Negative (No Plasid) 36ul Ampicillin (50ug/ml)


Wednesday (18.7.12)

Aim - Results from Transformation

Result: Table below indicates there was no growth for our cells, but that the controls worked. Also included is an image of each plate.

Samples Growth/No Growth
Plasmid pTOP No Growth
Control Positive (No Plasmid) Growth
Negative (No Plasid) No Growth


Wednesday (18.7.12)

Aim - Testing the transformation protocols: In light of the problems with cell competency, we decided to test whether the choice of protocol was contributing to the poor transformation results – especially as it would require another week to set up another cell line. We used a known competent cell line, and tested its competency against both of our cell lines, for Transformation Protocol 1 vs Transformation Protocol 2. Each sample was plated on an Ampicillin positive Agar and incubated overnight.

(LOGO) Transformation - Protocol 1

(LOGO) Transformation - Protocol 2

Step ?: The table below describes the plates necessary for this investigation, each of which shoud carry Ampicillin antibiotic at a concentration of 50ug/ml.

Plates
Protocol 1 W3100 – Original
W3100 – Reinvigorated
W3100 – Known Competent
Protocol 2 W3100 – Original
W3100 – Reinvigorated
W3100 – Known Competent


Thursday 19.7.12

Aim - Results from Transformation of pTop

Results: Table below indicates there was significant growth from the Original W3100 cells, but not from other cell lines.


6.3

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