Team:Goettingen/week21-2

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#2 Speed Improvement - 21st week

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V09_17

V09_17_1 Preparative double digestion of flhDC, FliC, 18K-RFP and pSB1C3 followed by ligation

Experiment:
In order to clone the flhDC, FliC and 18K-RFP constructs into pSB1C3, all components were digested with EcoRI and PstI according to the protocol. The restriction products were subsequently separated on a gel in order to verify the digestions success and purify the desired fragments. After measuring the concentration of the purified fragments, an overnight ligation was initiated.

V09_17_2 Preparation of over night cultures

Experiment: We prepared overnight cultures of different promoter-gene constructs in two different E. coli strains (MG1655 & BL21) in order to test wether the motility is influenced. The following constructs were used:

- 20E_flhDC
- 18C_flhDC
- 20I_flhdc
- 20E_motA
- 18C_motA
- 20E_motB
- 18C_motB

V09_18

V09_18_1 Motility Assays

Experiment: For the motility assay the over night cultures, with different constructs, were spun down and dropped onto M9- or 1% trypton-agar. The exact protocol you can find HERE
Observation and Results:
Text

V09_18_2 Preparative double digestion of MotA, MotB, yhjH, 18C-RFP, 20E-RFP and 20I-RFP followed by ligation

Experiment:
TEXT

V09_18_3 Chemical transformation of different plasmid-gene constructs into E. coli (DH10B) or (MG1655)

Experiment:
Chemical transformation was done according to the standard protocol. The following constructs were used:
- pSB1C3-FliC into E. coli (DH10B)
pSB1C3-flhDC into E. coli (DH10B)
PSB1C3-RFP into E. coli (MG1655)

V09_18

V09_18_1 Motility Assays

Experiment: For the motility assay the over night cultures, with different constructs, were spun down and dropped onto M9- or 1% trypton-agar. The exact protocol you can find HERE
Observation and Results:
Text

V09_18_2 Preparative double digestion of MotA, MotB, yhjH, 18C-RFP, 20E-RFP and 20I-RFP followed by ligation

Experiment:
In order to test the influence of the different genes, we cloned the genes behind constitutive promoters with various promoter strength. MotA, MotB and yhjH were digested with XbaI and PstI. The psB1C3-Promoter backbones were digested with SpeI and PstI. Afterwards, the fragments of the desired size were cut out and purified via gel-extraction kit. Finally, over night ligation was started.

V09_18_3 Chemical transformation of different plasmid-gene constructs into E. coli (DH10B) or (MG1655)

Experiment:
Chemical transformation was done according to the standard protocol. The following constructs were used:
- pSB1C3-FliC into E. coli (DH10B)
pSB1C3-flhDC into E. coli (DH10B)
PSB1C3-RFP into E. coli (MG1655)

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@@ Line 534: / Line 534: @@
 <ul>
 <li>Experiment: <br>
-In order to clone the <i>flhDC</i>, <i>FliC</i> and 18K-<i>RFP</i> constructs into pSB1C3,  all components were digested with EcoRI and PstI according to the <a href="https://2012.igem.org/Team:Goettingen/Project/Methods">protocol</a>. The restriction products were subsequently separated on a gel in order to verify the digestions success and purify the desired fragments. After measuring the concentration of the purified fragments, an overnight ligation was initiated .<br>
+In order to clone the <i>flhDC</i>, <i>FliC</i> and 18K-<i>RFP</i> constructs into pSB1C3,  all components were digested with EcoRI and PstI according to the <a href="https://2012.igem.org/Team:Goettingen/Project/Methods">protocol</a>. The restriction products were subsequently separated on a gel in order to verify the digestions success and purify the desired fragments. After measuring the concentration of the purified fragments, an overnight ligation was initiated.<br>
 </ul>
 <br>
 <b>V09_17_2 Preparation of over night cultures</b><br>
           <ul>
-<li>Experiment: We prepared overnight cultures of different promoter-gene constructs in two different E. coli strains (MG1655 & BL21) in order to test wether the motolity is influenced. The following constructs were used: <br>
+<li>Experiment: We prepared overnight cultures of different promoter-gene constructs in two different E. coli strains (MG1655 & BL21) in order to test wether the motility is influenced. The following constructs were used: <br>
 <br>
 - 20E_<i>flhDC</i> <br>