LOCUS       Exported                6129 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pCyPet-His
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6129)
  AUTHORS   Nguyen AW, Daugherty PS
  TITLE     Evolutionary optimization of fluorescent proteins for intracellular 
            FRET.
  JOURNAL   Nat Biotechnol. 2005 Mar . 23(3):355-60.
  PUBMED    15696158
REFERENCE   2  (bases 1 to 6129)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6129
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(73..90)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      243..329
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      421..448
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        467..558
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      1016..1471
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1103..1122)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     CDS             complement(2026..2685)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2600..2619
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(2686..2788)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(3180..3197)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3314..3859)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(3394..3413)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3931..3950
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             complement(4130..5008)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        5035..5319
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     5242..5261
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             5377..6090
                     /codon_start=1
                     /product="cyan fluorescent protein optimized for FRET with 
                     YPet (Nguyen and Daugherty, 2005)"
                     /label=CyPet
                     /translation="MSKGEELFGGIVPILVELEGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSVMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIKA
                     NFKARHNITDGSVQLADHYQQNTPIGDGPVILPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             6091..6108
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
ORIGIN
        1 ccggggatcc tctagagtcg acctgcaggc atgcaagctt ggctgttttg gcggatgaga
       61 gaagattttc agcctgatac agattaaatc agaacgcaga agcggtctga taaaacagaa
      121 tttgcctggc ggcagtagcg cggtggtccc acctgacccc atgccgaact cagaagtgaa
      181 acgccgtagc gccgatggta gtgtggggtc tccccatgcg agagtaggga actgccaggc
      241 atcaaataaa acgaaaggct cagtcgaaag actgggcctt tcgttttatc tgttgtttgt
      301 cggtgaacgc tctcctgagt aggacaaatc cgccgggagc ggatttgaac gttgcgaagc
      361 aacggcccgg agggtggcgg gcaggacgcc cgccataaac tgccaggcat caaattaagc
      421 agaaggccat cctgacggat ggcctttttg cgtttctaca aactcttttg tttatttttc
      481 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
      541 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
      601 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
      661 gaagatcagt tgggtgcagc aaactattaa ctggcgaact acttactcta gcttcccggc
      721 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
      781 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
      841 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
      901 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
      961 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttacgcg
     1021 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     1081 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     1141 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     1201 ttacggcacc tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag tgggccatcg
     1261 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     1321 ttgttccaaa cttgaacaac actcaaccct atctcgggct attcttttga tttataaggg
     1381 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     1441 aattttaaca aaatattaac gtttacaatt taaaaggatc taggtgaaga tcctttttga
     1501 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     1561 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     1621 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     1681 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta
     1741 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     1801 aatcctgtta ccagtcaggc atttgagaag cacacggtca cactgcttcc ggtagtcaat
     1861 aaaccggtaa accagcaata gacataagcg gctatttaac gaccctgccc tgaaccgacg
     1921 accgggtcga atttgctttc gaatttctgc cattcatccg cttattatca cttattcagg
     1981 cgtagcacca ggcgtttaag ggcaccaata actgccttaa aaaaattacg ccccgccctg
     2041 ccactcatcg cagtactgtt gtaattcatt aagcattctg ccgacatgga agccatcaca
     2101 gacggcatga tgaacctgaa tcgccagcgg catcagcacc ttgtcgcctt gcgtataata
     2161 tttgcccatg gtgaaaacgg gggcgaagaa gttgtccata ttggccacgt ttaaatcaaa
     2221 actggtgaaa ctcacccagg gattggctga gacgaaaaac atattctcaa taaacccttt
     2281 agggaaatag gccaggtttt caccgtaaca cgccacatct tgcgaatata tgtgtagaaa
     2341 ctgccggaaa tcgtcgtggt attcactcca gagcgatgaa aacgtttcag tttgctcatg
     2401 gaaaacggtg taacaagggt gaacactatc ccatatcacc agctcaccgt ctttcattgc
     2461 catacggaat tccggatgag cattcatcag gcgggcaaga atgtgaataa aggccggata
     2521 aaacttgtgc ttatttttct ttacggtctt taaaaaggcc gtaatatcca gctgaacggt
     2581 ctggttatag gtacattgag caactgactg aaatgcctca aaatgttctt tacgatgcca
     2641 ttgggatata tcaacggtgg tatatccagt gatttttttc tccattttag cttccttagc
     2701 tcctgaaaat ctcgataact caaaaaatac gcccggtagt gatcttattt cattatggtg
     2761 aaagttggaa cctcttacgt gccgatcaac gtctcatttt cgccaaaagt tggcccaggg
     2821 cttcccggta tcaacaggga caccaggatt tatttattct gcgaagtgat cttccgtcac
     2881 aggtatttat tcggcgcaaa gtgcgtcggg tgatgctgcc aacttactga tttagtgtat
     2941 gatggtgttt ttgaggtgct ccagtggctt ctgtttctat cagctgtccc tcctgttcag
     3001 ctactgacgg ggtggtgcgt aacggcaaaa gcaccgccgg acatcagcgc tagcggagtg
     3061 tatactggct tactatgttg gcactgatga gggtgtcagt gaagtgcttc atgtggcagg
     3121 agaaaaaagg ctgcaccggt gcgtcagcag aatatgtgat acaggatata ttccgcttcc
     3181 tcgctcactg actcgctacg ctcggtcgtt cgactgcggc gagcggaaat ggcttacgaa
     3241 cggggcggag atttcctgga agatgccagg aagatactta acagggaagt gagagggccg
     3301 cggcaaagcc gtttttccat aggctccgcc cccctgacaa gcatcacgaa atctgacgct
     3361 caaatcagtg gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggcg
     3421 gctccctcgt gcgctctcct gttcctgcct ttcggtttac cggtgtcatt ccgctgttat
     3481 ggccgcgttt gtctcattcc acgcctgaca ctcagttccg ggtaggcagt tcgctccaag
     3541 ctggactgta tgcacgaacc ccccgttcag tccgaccgct gcgccttatc cggtaactat
     3601 cgtcttgagt ccaacccgga aagacatgca aaagcaccac tggcagcagc cactggtaat
     3661 tgatttagag gagttagtct tgaagtcatg cgccggttaa ggctaaactg aaaggacaag
     3721 ttttggtgac tgcgctcctc caagccagtt acctcggttc aaagagttgg tagctcagag
     3781 aaccttcgaa aaaccgccct gcaaggcggt tttttcgttt tcagagcaag agattacgcg
     3841 cagaccaaaa cgatctcaag aagatcatct tattaatcag ataaaatatt tgctcatgag
     3901 cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg gcgatatagg cgccagcaac
     3961 cgcacctgtg gcgccggtga tgccggccac gatgcgtccg gcgtagagga tctgctcatg
     4021 tttgacagct tatcatcgat gcataatgtg cctgtcaaat ggacgaagca gggattctgc
     4081 aaaccctatg ctactccgtc aagccgtcaa ttgtctgatt cgttaccaat tatgacaact
     4141 tgacggctac atcattcact ttttcttcac aaccggcacg gaactcgctc gggctggccc
     4201 cggtgcattt tttaaatacc cgcgagaaat agagttgatc gtcaaaacca acattgcgac
     4261 cgacggtggc gataggcatc cgggtggtgc tcaaaagcag cttcgcctgg ctgatacgtt
     4321 ggtcctcgcg ccagcttaag acgctaatcc ctaactgctg gcggaaaaga tgtgacagac
     4381 gcgacggcga caagcaaaca tgctgtgcga cgctggcgat atcaaaattg ctgtctgcca
     4441 ggtgatcgct gatgtactga caagcctcgc gtacccgatt atccatcggt ggatggagcg
     4501 actcgttaat cgcttccatg cgccgcagta acaattgctc aagcagattt atcgccagca
     4561 gctccgaata gcgcccttcc ccttgcccgg cgttaatgat ttgcccaaac aggtcgctga
     4621 aatgcggctg gtgcgcttca tccgggcgaa agaaccccgt attggcaaat attgacggcc
     4681 agttaagcca ttcatgccag taggcgcgcg gacgaaagta aacccactgg tgataccatt
     4741 cgcgagcctc cggatgacga ccgtagtgat gaatctctcc tggcgggaac agcaaaatat
     4801 cacccggtcg gcaaacaaat tctcgtccct gatttttcac caccccctga ccgcgaatgg
     4861 tgagattgag aatataacct ttcattccca gcggtcggtc gataaaaaaa tcgagataac
     4921 cgttggcctc aatcggcgtt aaacccgcca ccagatgggc attaaacgag tatcccggca
     4981 gcaggggatc attttgcgct tcagccatac ttttcatact cccgccattc agagaagaaa
     5041 ccaattgtcc atattgcatc agacattgcc gtcactgcgt cttttactgg ctcttctcgc
     5101 taaccaaacc ggtaaccccg cttattaaaa gcattctgta acaaagcggg accaaagcca
     5161 tgacaaaaac gcgtaacaaa agtgtctata atcacggcag aaaagtccac attgattatt
     5221 tgcacggcgt cacactttgc tatgccatag catttttatc cataagatta gcggatccta
     5281 cctgacgctt tttatcgcaa ctctctactg tttctccata cccgtttttt tgggctagcg
     5341 aattcgagct cggccacgaa ggccaggagt gaaacgatgt ctaaaggtga agaattattc
     5401 ggcggtatcg tcccaatttt agttgaatta gagggtgatg ttaatggtca caaattttct
     5461 gtctccggtg aaggtgaagg tgatgctacg tacggtaaat tgaccttaaa atttatttgt
     5521 actactggta aattgccagt tccatggcca accttagtca ctactctgac ttggggtgtt
     5581 caatgttttt ctagataccc agatcatatg aaacaacatg actttttcaa gtctgtcatg
     5641 ccagaaggtt atgttcaaga aagaactatt tttttcaaag atgacggtaa ctacaagacc
     5701 agagctgaag tcaagtttga aggtgatacc ttagttaata gaatcgaatt aaaaggtatt
     5761 gattttaaag aagatggtaa cattttaggt cacaaattgg aatacaacta tatctctcac
     5821 aatgtttaca tcaccgctga caaacaaaag aatggtatca aagctaactt caaagccaga
     5881 cacaacatta ccgatggttc tgttcaatta gctgaccatt atcaacaaaa tactccaatt
     5941 ggtgatggtc cagtcatctt gccagacaac cattacttat ccactcaatc tgccttatct
     6001 aaagatccaa acgaaaagag agaccacatg gtcttgctcg aatttgttac tgctgctggt
     6061 attacccatg gtatggatga attgtacaaa caccaccacc accaccacta aggccaaggt
     6121 ggccggtac
//
