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Supplementary Note Vectors are available upon request by non-commercial users. GFP 1–10 and GFP 11 can also be prepared by primer-directed mutation of GFPs on commercially available plasmids or by ordering coding sequences from gene synthesis companies. Amino acid and sequence of the GFP 11 cassette: SSLKRRKIPMGSSHHHHHHSSGLVPRGSHM*(frame shift +1) *LIGSDGGSGGGSTSRDHMVLHEYVNAAGIT*GT*LEHHHHHH*DPAANKARKEAELAAA TAEQ*LA*PLEA DNA sequence of the GFP 11 cassette 0421007 TAGAGATACTGAGCACATCAGCAGGACGCACTGACCGAGTTCATTAAAGAGGAGAAAGA TACCATGGGCAGCAGCCATCATCATCATCATCACAGCAGCGGCCTGGTGCCGCGCGGCA GCCATATGTAATTAATTAATTGGATCCGATGGAGGGTCTGGTGGCGGATCAACAAGTCG TGACCACATGGTCCTTCATGAGTACGTAAATGCTGCTGGGATTACATAAGGTACCTAAC TCGAGCACCACCACCACCACCACTGAGATCCGGCTGCTAACAAAGCCCGAAAGGAAGCT GAGTTGGCTGCTGCCACCGCTGAGCAATAACTAGCATAACCTCTAGAGGCCTC 02129811 Amino acid sequence of the GFP 1-10 cassette DRDLDPAKLIRLTIHMGGTSMSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATIG KLTLKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISF KDDGKYKTRAVVKFEGDTLVNRIELKGTDFKEDGNILGHKLEYNFNSHNVYITADKQKNG IKANFTVRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQTVLSKDPNEK*GTLEH HHHHH*DPAANKARKEAELAAATAEQ*LA*PLEA DNA sequence of the GFP 1-10 cassette 01079936 AGGATCGAGATCTCGATCCCGCGAAATTAATACGACTCACTATACATATGGGTGGCACTA GTATGAGCAAAGGAGAAGAACTTTTCACTGGAGTTGTCCCAATTCTTGTTGAATTAGATG GTGATGTTAATGGGCACAAATTTTCTGTCAGAGGAGAGGGTGAAGGTGATGCTACAATCG GAAAACTCACCCTTAAATTTATTTGCACTACTGGAAAACTACCTGTTCCATGGCCAACAC TTGTCACTACTCTGACCTATGGTGTTCAATGCTTTTCCCGTTATCCGGATCACATGAAAA GGCATGACTTTTTCAAGAGTGCCATGCCCGAAGGTTATGTACAGGAACGCACTATATCTT TCAAAGATGACGGGAAATACAAGACGCGTGCTGTAGTCAAGTTTGAAGGTGATACCCTTG TTAATCGTATCGAGTTAAAGGGTACTGATTTTAAAGAAGATGGAAACATTCTCGGACACA AACTCGAGTACAACTTTAACTCACACAATGTATACATCACGGCAGACAAACAAAAGAATG GAATCAAAGCTAACTTCACAGTTCGCCACAACGTTGAAGATGGTTCCGTTCAACTAGCAG ACCATTATCAACAAAATACTCCAATTGGCGATGGCCCTGTCCTTTTACCAGACAACCATT ACCTGTCGACACAAACTGTCCTTTCGAAAGATCCCAACGAAAAGTAAGGTACCCTCGAG CACCACCACCACCACCACTGAGATCCGGCTGCTAACAAAGCCCGAAAGGAAGCTGAGTT GGCTGCTGCCACCGCTGAGCAATAACTAGCATAACCTCTAGAGGCCTC 02129811 Vector primers to reamplify the split GFP fragments cassette Top primer for pTET GFP11 plasmid: 0421007 5’-TAGAGATACTGAGCACATCAGCAGGACGCACTGACC-3’ Bottom primer for pTET GFP11 and pET GFP1-10 plasmids: 02129811 5’-GAGGCCTCTAGAGGTTATGCTAGTTATTGC-3’ Top primer for pET GFP1-10 plasmid: 01079936 5’-AGGATCGAGATCTCGATCCCGCGAAATTAATAC-3’ The GFP 11 cassette can be amplified from pTET plasmid using 0421007 and 02129811 primers. The restriction sites for cloning insert proteins is indicated in bold in the DNA sequence. A frameshift stuffer with three translational stops (one in each frame) prevents false positives from a self-religated plasmid. A 6His tag followed by a thrombin cleavage site is located at the N terminus. The initiator methionine is indicated in bold in the amino-acid sequence of both GFP 11 and GFP 1-10 cassettes. The GFP 1-10 cassette can be amplified using 01079936 and 02129811 primers.
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