Bio Sketch The bacterial sketch pad Yves Wang
Bio. Sketch The bacterial sketch pad. Yves Wang, Jennifer Gao, Yin Li Chris Doucette, Thomas Noriega, Hing Eng Group Meeting 2005 -07 -05
Bio. Sketch Harvard i. GEM 2005 Talk outline What has been done with the Collins constructs What has been done with the Bio. Bricks What’s in store for this week 2
Bio. Sketch Harvard i. GEM 2005 Collins (Yin and Jenny) 3
Bio. Sketch Harvard i. GEM 2005 What has been done with Biobricks Pairwise assembly: QPI digestions successful with modified protocol: Pl + QPILac QPIl + YFP QPI 434 + YFP Completed first two rounds and started third round of ligations to build QPIs from scratch Performing ligations in duplicate to decrease chances of setbacks Sequenced successful ts (mut 265) mutations in Lac. I Have sent Lac. I ts (mut 241) and both QPILac. I(ts) mutants (m 241 and m 265) for sequencing Outsourced 2 different ts, ind- versions of lc. I. (“m. JP 3” based on Japanese patent Genbank #E 07700, and “m 8572” based on c. I 857 sequence found in HENDRIX Lambda II book). 4
Bio. Sketch Harvard i. GEM 2005 Successful Ligations: Ligations m. Cherry + Terminator RBS + 434 RBS + lc. I RBS + m. Cherry-Terminator RBS + Venus-Terminator Digestions ready for ligation: Plambda. CI P 434 Plac. I(hybrid) 434 lc. I RBS-434 RBS-lc. I RBS-m. Cherry-Terminator RBS-Venus-Terminator QPIlac. I QPIl QPI 434 5
Bio. Sketch Harvard i. GEM 2005 Summary: What we have done Collins. Mod: (? ) Bio. Bricks: Pairwise assembly m. Cherry + T RBS + 434 RBS + lc. I RBS + m. Cherry-Terminator RBS + Venus-Terminator QPI digest problem solved; ligations still in testing Mutations and Sequencing We have a sequenced Bb-Lac. I m 265 We are sequencing Lac. I m 241 and QPI-lac. I m 241 and m 265 Introducing m. JP 3 and m 8572 into Lambda c. I and QPI-Lambda c. I 6
Bio. Sketch Harvard i. GEM 2005 This week with Collins 7
Bio. Sketch Harvard i. GEM 2005 This week with Biobricks More Pairwise Assembly First Round Plac. I(hyb) + RBS-l. CI Pl + RBS-434 RBS + Lac. Its(265) All promoters + m. Cherry All promoters + Venus Second Round Terminator + P 434 -m. Cherry Terminator + P 434 -Venus Terminator + Pl -m. Cherry Terminator + Pl -Venus Sequence c. I 857 mutations QPIl + m. Cherry QPI 434 + m. Cherry Pl + QPIlac. I Pl-QPIlac. I + QPIl-m. Cherry Pl-QPIlac. I + m. Cherry Plac. I + QPIl-m. Cherry Pl + QPI 434 -m. Cherry Test microscopy with Promoter-Reporter constructs Are m. Cherry-expressing colonies red enough? Make Tsien lab bacterial m. Cherry into Bio. Brick 8
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