LOCUS Exported 5302 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS pRSET FLII32Pglu-2.2m SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5302) AUTHORS Deuschle K, Okumoto S, Fehr M, Looger LL, Kozhukh L, Frommer WB TITLE Construction and optimization of a family of genetically encoded metabolite sensors by semirational protein engineering. JOURNAL Protein Sci. 2005 Sep . 14(9):2304-14. PUBMED 16131659 REFERENCE 2 (bases 1 to 5302) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5302 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 19..38 /label=T7 /note="T7 promoter, forward primer" promoter 19..37 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" RBS 67..89 /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" CDS 109..126 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" CDS 130..162 /codon_start=1 /product="leader peptide from bacteriophage T7 gene 10" /label=T7 tag (gene 10 leader) /note="promotes efficient translation in E. coli" /translation="MASMTGGQQMG" CDS 166..189 /codon_start=1 /product="Xpress(TM) epitope tag, including an enterokinase recognition and cleavage site" /label=Xpress(TM) tag /translation="DLYDDDDK" CDS 202..918 /codon_start=1 /product="enhanced YFP" /label=EYFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 1039..1752 /codon_start=1 /product="enhanced CFP" /label=ECFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELY" primer_bind complement(2648..2666) /label=T7 Term /note="T7 terminator, reverse primer" terminator 2662..2709 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" rep_origin 2806..3261 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2893..2912) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3103..3124 /label=F1ori-F /note="F1 origin, forward primer" promoter 3342..3446 /gene="bla" /label=AmpR promoter CDS 3447..4307 /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCDTLLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDESDTTMPVAMPTTLRKLLTGELLTLASRQQLID*MEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKRSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" rep_origin 4478..5066 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 4967..4986 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 5220..5237 /label=L4440 /note="L4440 vector, forward primer" ORIGIN 1 atctcgatcc cgcgaaatta atacgactca ctatagggag accacaacgg tttccctcta 61 gataattttg tttaacttta agaaggagat atacatatgc ggggttctca tcatcatcat 121 catcatggta tggctagcat gactggtgga cagcaaatgg gtcgggatct gtacgacgat 181 gacgataagg atccgggccg catggtgagc aagggcgagg agctgttcac cggggtggtg 241 cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag 301 ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag 361 ctgcccgtgc cctggcccac cctcgtgacc accttcggct acggcctgca gtgcttcgcc 421 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac 481 gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg 541 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag 601 gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc 661 atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag 721 gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc 781 gtgctgctgc ccgacaacca ctacctgagc taccagtccg ccctgagcaa agaccccaac 841 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc 901 atggacgagc tgtacaaggg tggtaccgga ggcgccgctg atactcgcat tggtgtaaca 961 atctataagt acgacgataa cgcgatgtct gtagtgcgca aggctattga gcaagatgcg 1021 aaagccgcgt cggctggtat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc 1081 atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc 1141 gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg 1201 cccgtgccct ggcccaccct cgtgaccacc ctgacctggg gcgtgcagtg cttcagccgc 1261 taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc 1321 caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag 1381 ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac 1441 ggcaacatcc tggggcacaa gctggagtac aactacatca gccacaacgt ctatatcacc 1501 gccgacaagc agaagaacgg catcaaggcc aacttcaaga tccgccacaa catcgaggac 1561 ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg 1621 ctgctgcccg acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag 1681 aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg 1741 gacgagctgt acggtagccg agttcagctg ctgatgaatg attctcagaa tgaccagtcc 1801 aagcagaacg atcagatcga cgtattgctg gcgaaagggg tgaaggcact ggcaatcaac 1861 ctggttgacc cggcagctgc gggtacggtg attgagaaag cgcgtgggca aaacgtgccg 1921 gtggttttct tcaacaaaga accgtctcgt aaggcgctgg atagctacga caaagcctac 1981 tacgttggca ctgactccaa agagtccggc attattcaag gcgatttgat tgctaaacac 2041 tgggcggcga atcagggttg ggatctgaac aaagacggtc agattcagtt cgtactgctg 2101 aaaggtgaac cgggccatcc ggatgcagaa gcacgtacca cttacgtgat taaagaattg 2161 aacgataaag gcatcaaaac tgaacagtta cagttagata ccgcaatgtg ggacaccgct 2221 caggcgaaag ataagatgga cgcctggctg tctggcccga acgccaacaa aatcgaagtg 2281 gttatcgcca acaacgatgc gatggcaatg ggcgcggttg aagcgctgaa agcacacaac 2341 aagtccagca ttccggtgtt tggcgtcgat gcgctgccag aagcgctggc gctggtgaaa 2401 tccggtgcac tggcgggcac cgtactgaac gatgctaaca accaggcgaa agcgaccttt 2461 gatctggcga aaaacctggc cgatggtaaa ggtgcggctg atggcaccaa ctggaaaatc 2521 gacaacaaag tggtccgcgt accttatgtt ggcgtagata aagacaacct ggctgaattc 2581 agcaagaaat aaaagcttga tccggctgct aacaaagccc gaaaggaagc tgagttggct 2641 gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg 2701 ggttttttgc tgaaaggagg aactatatcc ggatctggcg taatagcgaa gaggcccgca 2761 ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atgggacgcg ccctgtagcg 2821 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg 2881 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc 2941 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagagct ttacggcacc 3001 tcgaccgcaa aaaacttgat ttgggtgatg gttcacgtag tgggccatcg ccctgataga 3061 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa 3121 ctggaacaac actcaaccct atcgcggtct attcttttga tttataaggg attttgccga 3181 tttcggccta ttggttaaaa aatgagctga tttaacaaat atttaacgcg aattttaaca 3241 aaatattaac gtttacaatt tcgcctgatg cggtattttc tccttacgca tctgtgcggt 3301 atttcacacc gcatacaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt 3361 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc 3421 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc 3481 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa 3541 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg 3601 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag 3661 ttctgctatg tgatacacta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc 3721 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta 3781 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg 3841 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca 3901 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac 3961 caaacgacga gagtgacacc acgatgcctg tagcaatgcc aacaacgttg cgcaaactat 4021 taactggcga actacttact ctagcttccc ggcaacaatt aatagactga atggaggcgg 4081 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata 4141 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta 4201 agcgctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa 4261 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag 4321 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg 4381 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact 4441 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg 4501 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc 4561 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata 4621 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta 4681 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc 4741 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg 4801 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac 4861 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg 4921 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt 4981 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct 5041 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg 5101 gcttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata 5161 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca 5221 gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc 5281 gttggccgat tcattaatgc ag //