IGEM:Harvard/2007/Gels

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(6/27 plasmid gel, performed by Stephanie)
Current revision (10:39, 30 July 2007) (view source)
(7/09 Egel, Alex)
 
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[[Image:Gelpostpcrplasmid0627.jpg|thumb|left|250px]]
[[Image:Gelpostpcrplasmid0627.jpg|thumb|left|250px]]
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<br><br><br>
 
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Description: Gel using Lpp+OmpA1+pet29 and LppOmpA2+pet29 <br>
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Description: Gel using Lpp+OmpA1+pet29 and LppOmpA2+pet29  
Explanation:  
Explanation:  
For further information concerning amounts of plasmid in sample, please refer to Stephanie's notebook page (under Stephanie's personal page, shlo/notebook)
For further information concerning amounts of plasmid in sample, please refer to Stephanie's notebook page (under Stephanie's personal page, shlo/notebook)
Wells are as follows:<br>
Wells are as follows:<br>
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#Well 1: 10uL ladder of 1kB plus, from Invitrogen
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Well 1: 10uL ladder of 1kB plus, from Invitrogen
<i> Note: this ladder cannot be used to quantify - perhaps a reason we should just use 1kB ladder, rather than 1kB+, in the future</i><br>
<i> Note: this ladder cannot be used to quantify - perhaps a reason we should just use 1kB ladder, rather than 1kB+, in the future</i><br>
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#Well 2: 1uL OmpA1plasmid (diluted to 20uL)
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Well 2: 1uL OmpA1plasmid (diluted to 20uL)<br>
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#Well 3: 3uL OmpA1plasmid (diluted to 20)
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Well 3: 3uL OmpA1plasmid (diluted to 20)<br>
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#Well 4: 1uL OmpA2plasmid (diluted to 20)
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Well 4: 1uL OmpA2plasmid (diluted to 20)<br>
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#Well 5: 3uL OmpA2plasmid (diluted to 20)
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Well 5: 3uL OmpA2plasmid (diluted to 20)<br><br>
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== 6/27 protein denaturation electrophoresis gel ==
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[[Image:1_protGel_Jul7.jpg|thumb|left|250px]]<br>
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<br>Description: 6/27 denaturation electrophoresis gel <br>
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The rightmost lane is the ladder
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lane 1 Ladder <br>
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lane 2 OmpA1 + strep before induction with IPTG<br>
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lane 3 OmpA1 + his6 before induction<br>
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lane 4 OmpA1 + strep after induction<br>
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lane 5 OmpA1 + his6 after induction<br>
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lane 6 OmpA1 + his6 after induction<br>
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lane 7 OmpA1 + strep after induction<br>
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lane 8 OmpA1 + his6 before induction<br>
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lane 9 OmpA1 + strep before induction<br><br>
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''Note that there were 2 copies of each sample: samples in lanes 2 to 5 were denatured for 2 minutes as per protocol while samples in lanes 6 to 9 were denatured for a 10 minutes, Sammy S.''
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== 7/05 protein gel, Kevin and Sammy ==
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[[Image: 2007-07-06protGel2.jpg|thumb|left|300px]]<br>
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'''Gel 2: induced immediately'''<br>
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Lane 1: Ladder<br>
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Lane 2: Strep No IPTG<br>
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Lane 3: Strep denatured after 30 min<br>
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Lane 4: Strep denatured after 60 min<br>
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Lane 5: Strep denatured after 90 min<br>
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Lane 6: Strep denatured after 120 min <br>
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Lane 7: His  No IPTG<br>
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Lane 8: His denatured after 30 min <br>
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Lane 9: His denatured after 60 min <br>
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Lane 10: His denatured after 90 min <br><br><br><br>
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[[Image: 2007-07-06protGel1.jpg|thumb|left|350px]]<br>
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'''Gel 1'''<br>
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Lane 1: Ladder<br>
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Lane 2: His 0 min induction<br>
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Lane 3: His 30 min induction<br>
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Lane 4: His 60 min induction<br>
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Lane 5: His 90 min induction<br>
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Lane 6: His 120 min induction<br>
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Lane 7: Strep 0 min induction<br>
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Lane 8: Strep 30 min induction<br>
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Lane 9: Strep 60 min induction<br>
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Lane 10: Strep 90 min induction<br>
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Lane 11: Strep 120 min induction<br>
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Lane 12: His 0 min induction 120 min denature<br>
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 +
 
 +
== 7/09 Egel, Alex ==
 +
[[Image: mutagenesisgel.jpg|thumb|left|300px]]<br>
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'''Gel 2: induced immediately'''<br>
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Lane 1: Ladder<br>
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Lane 2: Undigested FecR (-) R1A<br>
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Lane 3: Pst1 Digested Fec R (-) R1A<br>
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Lane 4: Undigested FecR (-) R5B<br>
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Lane 5: Pst1 Digested Fec R (-) R5B<br>
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Lane 6: Pst1 Digested Fec R (+) <br>
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Lane 7: Undigested FecA (-) A1A<br>
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Lane 8: Pst1 Digested Fec A (-) A1A<br>
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Lane 9: Undigested FecR (-) A5B<br>
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Lane 10: Pst1 Digested Fec R (-) A5B<br>
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Lane 11: Pst1 Digested Fec R (+) A1B<br>
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Lane 12: Pst1 Digested Fec A (+) <br>
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<br>
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 +
== 7/26/07 Egel, Kevin and Mike ==
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[[Image: KevinMikeGel.jpg|thumb|left|300px]]<br>
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'''Gel: AIDA after PCR'''<br>
 +
 
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Lane 1: Ladder<br>
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Lane 2: AIDA1+his<br>
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Lane 3: AIDA1+strep<br>
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Result: We did not get ideal results, though we did get light expression where we expected.

Current revision

Contents

Week 0 gel


Description: Week 0 gel
Well 1 Ladder
Well 2 OmpA1 in PET
Well 3 OmpA2 in PET
Well 4 OmpA1 +his tag in PET
Well 5 OmpA1 +strep tag in PET
Well 6 OmpA2 +his tag in PET
Well 7 OmpA2 +strep tag in PET



6/20 gel

Description: 6/20 gel
Well 1 Ladder (10 kb)
Well 2 OmpA1, Nhe1, Pst1, Group 1 27.5 uL
Well 3 OmpA1, Nhe1, Pst1, Group 1 27.5 uL
Well 4 OmpA1, Nhe1, Pst1, Group 2 27.5 uL
Well 5 OmpA1, Nhe1, Pst1, Group 2 27.5 uL
Well 6 OmpA1, Nhe1, Pst1, Group 2 27.5 uL
Well 7 OmpA2, Nhe1, Pst1, Group 4 27.5 uL
Well 8 OmpA2, Nhe1, Pst1, Group 4 27.5 uL
Well 9 OmpA2, Nhe1, Pst1, Group 3 27.5 uL
Well 10 OmpA2, Nhe1, Pst1, Perry's 27.5 uL
Well 11 OmpA2, Nhe1, Pst1, Perry's
See Notebook page for gel extraction weights and values.

6/19 gel after excision


Description: 6/19 gel after excision

Well 1 Ladder
Well 2 OmpA1 in PET
Well 3 OmpA2 in PET
Well 4 OmpA1 +his tag in PET
Well 5 OmpA1 +strep tag in PET
Well 6 OmpA2 +his tag in PET
Well 7 OmpA2 +strep tag in PET



6/22 overexpression gel


Description: 6/22 overexpression gel
lane 1 Ladder
lane 2 OmpA1 + his6 non-induced
lane 3 OmpA1 + his6 induced
lane 4 OmpA1 + strep2 non-induced
lane 5 OmpA1 + strep2 induced
lane 6 OmpA2 + his6 non-induced
lane 7 OmpA2 + his6 induced
lane 8 OmpA2 + strep2 non-induced
lane 9 OmpA2 + strep2 induced

6/22 overexpression gel2, ladder contaminated




Description: Contaminated 6/22 overexpression gel

Explanation: Failed gel because of contamination in each lane by the ladder, which was mistakenly added.





6/27 plasmid gel, performed by Stephanie

Description: Gel using Lpp+OmpA1+pet29 and LppOmpA2+pet29

Explanation: For further information concerning amounts of plasmid in sample, please refer to Stephanie's notebook page (under Stephanie's personal page, shlo/notebook) Wells are as follows:
Well 1: 10uL ladder of 1kB plus, from Invitrogen Note: this ladder cannot be used to quantify - perhaps a reason we should just use 1kB ladder, rather than 1kB+, in the future
Well 2: 1uL OmpA1plasmid (diluted to 20uL)
Well 3: 3uL OmpA1plasmid (diluted to 20)
Well 4: 1uL OmpA2plasmid (diluted to 20)
Well 5: 3uL OmpA2plasmid (diluted to 20)

6/27 protein denaturation electrophoresis gel



Description: 6/27 denaturation electrophoresis gel

The rightmost lane is the ladder lane 1 Ladder
lane 2 OmpA1 + strep before induction with IPTG
lane 3 OmpA1 + his6 before induction
lane 4 OmpA1 + strep after induction
lane 5 OmpA1 + his6 after induction
lane 6 OmpA1 + his6 after induction
lane 7 OmpA1 + strep after induction
lane 8 OmpA1 + his6 before induction
lane 9 OmpA1 + strep before induction

Note that there were 2 copies of each sample: samples in lanes 2 to 5 were denatured for 2 minutes as per protocol while samples in lanes 6 to 9 were denatured for a 10 minutes, Sammy S.

7/05 protein gel, Kevin and Sammy


Gel 2: induced immediately
Lane 1: Ladder
Lane 2: Strep No IPTG
Lane 3: Strep denatured after 30 min
Lane 4: Strep denatured after 60 min
Lane 5: Strep denatured after 90 min
Lane 6: Strep denatured after 120 min
Lane 7: His No IPTG
Lane 8: His denatured after 30 min
Lane 9: His denatured after 60 min
Lane 10: His denatured after 90 min




Gel 1
Lane 1: Ladder
Lane 2: His 0 min induction
Lane 3: His 30 min induction
Lane 4: His 60 min induction
Lane 5: His 90 min induction
Lane 6: His 120 min induction
Lane 7: Strep 0 min induction
Lane 8: Strep 30 min induction
Lane 9: Strep 60 min induction
Lane 10: Strep 90 min induction
Lane 11: Strep 120 min induction
Lane 12: His 0 min induction 120 min denature


7/09 Egel, Alex


Gel 2: induced immediately
Lane 1: Ladder
Lane 2: Undigested FecR (-) R1A
Lane 3: Pst1 Digested Fec R (-) R1A
Lane 4: Undigested FecR (-) R5B
Lane 5: Pst1 Digested Fec R (-) R5B
Lane 6: Pst1 Digested Fec R (+)
Lane 7: Undigested FecA (-) A1A
Lane 8: Pst1 Digested Fec A (-) A1A
Lane 9: Undigested FecR (-) A5B
Lane 10: Pst1 Digested Fec R (-) A5B
Lane 11: Pst1 Digested Fec R (+) A1B
Lane 12: Pst1 Digested Fec A (+)

7/26/07 Egel, Kevin and Mike


Gel: AIDA after PCR

Lane 1: Ladder
Lane 2: AIDA1+his
Lane 3: AIDA1+strep

Result: We did not get ideal results, though we did get light expression where we expected.

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