Template:SBB12Project stdt1200
From OpenWetWare
Welcome to your project page!
I've given you two parts to make.
- Design oligos to make your part
- Write up a proper construction file on the wiki template associated with your part
- Enter your Features (REMEMBER: It's not a bug, it's a feature. We just don't know what it does yet), Oligos, Parts, and Plasmids into Clotho
Also, use the link above left to upload a picture of yourself, your name, and anything else you'd like, and rename the link from "Your Name Here" to your name.
Finally, you should create a notebook on the on the main class page of the wiki
Create a LexA extravaganza.
LexA Fusion Two-Hybrid System Construction File: Construction of w.t. LexA PCR srs177f/srs178r on MG1655 genome (640 bp, pcrprdct1) Digest pcrprdct1 (Eco/Bam, 624, L, pcrdig) Digest pBca9145-part (Eco/Bam, L, vectdig) Ligate pcrdig, vectdig (9145-Bsrs038) srs177f CCATAgaattcATGagatctatgaaagcgttaacggccaggc srs178r gctagGGATCCttacagccagtcgccgttgcg Construction of LexA(1-87) with nhe1 end PCR srs177f/srs188r on w.t. LexA (292 bp, pcrprdct1) Digest pcrprdct1 (Eco/Bam, 276 L, pcrdig) Digest pBca9145-part (Eco/Bam, L, vectdig) Ligate pcrdig, vectdig (9145-Bsrs044) srs188r catagGCTAGCtggttcaccggcagccacacg Constructin of LexA408 (PA40, NS41, AS42) SOEing w.t.: cca-aac-gcg codon optimized PNA->ASS: gct-tct-tct ** PCR ca998/srs187r on 9145-Bsrs038 (222 bp, A) PCR g00101/srs186f on 9145-Bsrs038 (609 bp, B) PCR ca998/g00101 on A+B (783 bp, pcrprdct) Digest pcrprdct (Eco/Bam, 624, L, pcrdig) Digest pBca9145-part (Eco/Bam, L, vectdig) Ligate pcrdig, vectdig (9145-Bsrs039) srs186f GCGTTTGGGGTTCCGTTCCgcttcttctGCTGAAGAACATCTGAAGGC srs187r GCCTTCAGATGTTCTTCAGCagaagaagcGGAACGGAACCCCAAACGC Wobble to create sulA promoter with cononical RBS Wobble srs189f/srs190r (104bp, sulA1) Wobble srs191f/srs192r (104 bp, sulA2) Digest sulA1 (Eco/Bam 87+10+6, L, dig1) Digest sulA2 (Eco/Bam 87+10+6, L, dig2) Digest 9525 (Eco/Bam, L, vectdig) Ligate sulA1+9525 (Bsrs040) Ligate sulA1+9525 (Bsrs041) Bsrs040 is w.t. sulA promoter {op+/op+} Bsrs041 is mutant sulA promoter {408/op+} w.t sulA promoter (op+/op+): srs189f CCATAgaattcATGagatctaatagggttgatctttgttgtcaCTGGATGTACtGTACATCC Srs190r CTGATggatccaatcaatccagcccctgtgagttactgtatGGATGTACaGTACATCCAG Hybrid sulA promoter (408/op+): Srs191f CCATAgaattcATGagatctaatagggttgatctttgttgtcaCTGGATGTACCGTACATCC Srs192r CTGATggatccaatcaatccagcccctgtgagttactgtatGGATGTACGGTACATCCAG