Haynes Lab:Notebook/Characterizing AHL quorum sensing homologs/2013/05/23: Difference between revisions

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(Autocreate 2013/05/23 Entry for Haynes_Lab:Notebook/Characterizing_AHL_quorum_sensing_homologs)
 
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==Entry title==
==Entry title==
* Insert content here...
Miniprepped colonies picked yesterday


{| {{table}}
|-
| Part number || 260 || 280 || 260/280 || ng/ul
|-
| F2620 miniprep 1 || 0.052 || 0.027 || 1.881 || 51.662
|-
| F2620 miniprep 2 || 0.024 || 0.014 || 1.679 || 23.941
|-
| J06702 miniprep 1 || 0.039 || 0.022 || 1.766 || 38.85
|-
| J06702 miniprep 2 || 0.057 || 0.032 || 1.782 || 56.501
|-
| F2620-E0240 miniprep || 0.046 || 0.026 || 1.794 || 45.856
|}
<br>
Want to build F2620-J06702. Will try having F2620 as a front insert and J06702 as the backbone as well as F2620 as the backbone and J06702 as the rear insert.


Digests:
{| {{table}}
|-
| Number || Part number || Enzymes || Insert or Bb || Expected band sizes || Extracted band
|-
| 1|| J06702  || X/P || Rear insert || 869 + 2070 || 869
|-
| 2||J06702  || X/E|| Backbone || ~2900* || ~2900
|-
| 3||F2620  || S/P|| Backbone  || ~3100* || ~3100
|-
| 4||F2620 || E/S|| Front insert || 1061 + 2070 || 1061
|-
| 5||F2620-E0240  || X/P|| N/A || 1920 + 2070 || N/A
|}
*Not totally sure what the backbone length is.
Because of the low mini prep yields for 1-4, I used more DNA for digests 1-4:
{| {{table}}
|-
|Plasmid||25ul
|-
|MM||3ul
|-
|Enzyme 1||1ul
|-
|Enzyme 2||1ul
|-
|H20||0ul
|-
|Total|| 30ul
|}
For digest 5, I made a lower reaction volume because it's only to confirm:
{| {{table}}
|-
|Plasmid||10ul
|-
|MM||2ul
|-
|Enzyme 1||1ul
|-
|Enzyme 2||1ul
|-
|H20||6ul
|-
|Total|| 20ul
|}
<br>
Gel looked good but the J06702 vector is ~1kb longer than expected.
<br>
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__NOTOC__

Revision as of 12:28, 28 May 2013

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Entry title

Miniprepped colonies picked yesterday

Part number 260 280 260/280 ng/ul
F2620 miniprep 1 0.052 0.027 1.881 51.662
F2620 miniprep 2 0.024 0.014 1.679 23.941
J06702 miniprep 1 0.039 0.022 1.766 38.85
J06702 miniprep 2 0.057 0.032 1.782 56.501
F2620-E0240 miniprep 0.046 0.026 1.794 45.856


Want to build F2620-J06702. Will try having F2620 as a front insert and J06702 as the backbone as well as F2620 as the backbone and J06702 as the rear insert.

Digests:

Number Part number Enzymes Insert or Bb Expected band sizes Extracted band
1 J06702 X/P Rear insert 869 + 2070 869
2 J06702 X/E Backbone ~2900* ~2900
3 F2620 S/P Backbone ~3100* ~3100
4 F2620 E/S Front insert 1061 + 2070 1061
5 F2620-E0240 X/P N/A 1920 + 2070 N/A
  • Not totally sure what the backbone length is.

Because of the low mini prep yields for 1-4, I used more DNA for digests 1-4:

Plasmid 25ul
MM 3ul
Enzyme 1 1ul
Enzyme 2 1ul
H20 0ul
Total 30ul

For digest 5, I made a lower reaction volume because it's only to confirm:

Plasmid 10ul
MM 2ul
Enzyme 1 1ul
Enzyme 2 1ul
H20 6ul
Total 20ul


Gel looked good but the J06702 vector is ~1kb longer than expected.


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