Team:Grinnell/Notebook

From 2011.igem.org

(Difference between revisions)

Revision as of 22:16, 10 June 2011

Grinnell Menubar

Notebook

Week 1

PCR primers designed according to Biobrick specifications and ordered for rsaA C-terminal and WT esp. Because of the discrepancy in GC content between Staphylococcus epidermidis and Caulobacter, we also optimized the esp sequence for Caulobacter for later synthesis with a higher resulting GC content.

Prepared competent cells of E. coli Top10. (Protocol)

Streaked plates of Caulobacter, Staphylococcus aureus, and S. epidermidis.

Template DNA for PCR prepared from colonies using [http://www.bioventures.com/products.php?cid=10005| GeneReleaser]. (Protocol)

Week 2

Figure 1: PCR products on
DNA gel. Lane 1: ladder;
Lane 2: rsaA from liquid
culture Caulobacter; Lane 3:
rsaA from plate culture
Caulobacter; Lane 4: esp from
S. epidermidis.
Figure 2: Gel results for
transformation of ligations. Lane 1:
ladder; Lane 2: digested plasmid
with rsaA; Lane 3: digested plasmid
with rsaA and esp; Lanes 4-8:
digested plasmids from various
colonies with esp.

Performed PCR on esp and rsaA genes (protocol) and ran results on gel (protocol). Results showed successful amplification (figure 1).

Performed a transformation experiment (protocol) to test the efficiency of our competent cells make in Week 1. Results were successful with about a 0.7 x 10^7 CFU/μg of pUC19.

Purified PCR product (protocol) and digested esp with EcoRI, SpeI, and PstI; rsaA with EcoRI, XbaI, and PstI; and pSB1C3 with EcoRI and PstI. We then ligated esp, rsaA, and esp and rsaA to pSB1C3 and transformed into E. coli Top10. (Protocol)

Inoculated liquid cultures with colonies from transformations.

Performed digestion of results from transformation and ran on gel (figure 2).

Transformed BBa_K081005, a BioBrick part containing a gene for constitutive promotor and RBS, into E. coli Top10.

@@ Line 13: / Line 13: @@
 ===Week 2===
-<html><ul style="list-style:none; float:right;"><li><a href="https://2011.igem.org/File:20110606_PCRproduct.jpg"><img src="https://static.igem.org/mediawiki/2011/3/32/20110606_PCRproduct.jpg" alt="DNA gel from 20110606" height=200px/></a></li><li>Figure 1: PCR products on DNA gel.  Lane 1: ladder; Lane 2: <i>rsaA</i> from liquid culture <i>Caulobacter</i>; Lane 3: <i>rsaA</i> from plate culture <i>Caulobacter</i>; Lane 4: <i>esp</i> from <i>S. epidermidis</i>.</li><li><a href="https://2011.igem.org/File:20110610PlamidGel.jpg"><img src="https://static.igem.org/mediawiki/2011/a/a5/20110610PlamidGel.jpg" alt="Plasmid result gel for 20110610" height=200px/></a></li><li>Figure 2: Gel results for transformation of ligations. Lane 1: ladder; Lane 2: digested plasmid with <i>rsaA</i>; Lane 3: digested plasmid with <i>rsaA</i> and <i>esp</i>; Lanes 4-8: digested plasmids from various colonies with <i>esp</i>.</li></ul></html>
+<html><ul style="list-style:none; float:right;"><li><a href="https://2011.igem.org/File:20110606_PCRproduct.jpg"><img src="https://static.igem.org/mediawiki/2011/3/32/20110606_PCRproduct.jpg" alt="DNA gel from 20110606" height=200px/></a></li><li>Figure 1: PCR products on<br/>DNA gel.  Lane 1: ladder;<br/>Lane 2: <i>rsaA</i> from liquid<br/>culture <i>Caulobacter</i>; Lane 3:<br/><i>rsaA</i> from plate culture<br/><i>Caulobacter</i>; Lane 4: <i>esp</i> from<br/><i>S. epidermidis</i>.</li><li><a href="https://2011.igem.org/File:20110610PlamidGel.jpg"><img src="https://static.igem.org/mediawiki/2011/a/a5/20110610PlamidGel.jpg" alt="Plasmid result gel for 20110610" height=200px/></a></li><li>Figure 2: Gel results for<br/>transformation of ligations. Lane 1:<br/>ladder; Lane 2: digested plasmid<br/>with <i>rsaA</i>; Lane 3: digested plasmid<br/>with <i>rsaA</i> and <i>esp</i>; Lanes 4-8:<br/>digested plasmids from various<br/>colonies with <i>esp</i>.</li></ul></html>
 Performed PCR on esp and rsaA genes ([[Team:Grinnell/Notebook/Protocols#cPCR|protocol]]) and ran results on gel ([[Team:Grinnell/Notebook/Protocols#dGel|protocol]]).  Results showed successful amplification (figure 1).