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From 2014.igem.org

Plates from the Gibson assembly (pBbE1a-tet-CsgA) transformations grew many colonies! We picked 2 Turbo and 2 MACH1 colonies and inoculated in 5mL LB+Carb to shake up for mini-preps in the evening.

Minipreps:

→ Vector with ptet (we also have a streaked plate of this) → Vector with pBbE1a Backbone (we need to streak a plate of this for our stocks) → Picked colonies from the Gibson pBbE1a-tet-CsgA assembly

• We need to talk to Peter about getting a pET vector that we can miniprep/keep plates of
• We also want to talk to Peter about getting SSRA tags

We designed the following sequencing primers to confirm that our Gibson assembly worked and that the plasmid sequence was correct:

HBD5 GGAAGCTGTGGTATGGCTG HBD6 CAGCCATACCACAGCTTCC HBD7 GATACCGCTCGCCGCA HBD8 TGCGGCGAGCGGTATC HBD9 CGG CAA CCG AGC GTT CTG HBD10 CAG AAC GCT CGG TTG CCG HBD11 GGTTCCGCGCACATTTCC HBD12 GGAAATGTGCGCGGAACC

We sent the Gibson assembly plasmid for sequencing using the Bom_reverse primer, which is complementary to the vector’s backbone.