LOCUS       Exported                6595 bp ds-DNA     circular SYN 17-JAN-2024
DEFINITION  biosensor for cAMP in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    Pink Flamindo
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6595)
  AUTHORS   Harada K, Ito M, Wang X, Tanaka M, Wongso D, Konno A, Hirai H, 
            Hirase H, Tsuboi T, Kitaguchi T
  TITLE     Red fluorescent protein-based cAMP indicator applicable to 
            optogenetics and in vivo imaging.
  JOURNAL   Sci Rep. 2017 Aug 4;7(1):7351. doi: 10.1038/s41598-017-07820-6.
  PUBMED    28779099
REFERENCE   2  (bases 1 to 6595)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 17, 2024 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6595
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     1165..1184
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(1952..1969)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1958..2182
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2228..2656
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2315..2334)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2525..2546
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2665..2685)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2670..2999
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2850..2985
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2912..2931
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3066..3860
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3120..3139)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3730..3749
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4034..4155
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4071..4090)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4125..4144
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4204..4220)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4204..4220)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4217..4239)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4228..4244
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(4252..4282)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4297..4318
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4435..4452)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4606..5194)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4686..4705)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5365..6225)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5988..6007
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6226..6330)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6405..6424)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg
      661 gctagcgttt aaacgggccc tctagactcg agcggccgcc actgtgctgg atatctgcag
      721 aattccacca cactggacta gtggatccat gctggtgagc aagggcgagg aggataacat
      781 ggccatcatc aaggagttca tgcgcttcaa ggtgcacatg gagggctccg tgaacggcca
      841 cgagttcgag atcgagggcg agggcgaggg ccgcccctac gaggcctttc agaccgctaa
      901 gctgaaggtg accaagggtg gccccctgcc cttcgcctgg gacatcctgt cccctcagtt
      961 catgtacggc tccaaggcct acattaagca cccagccgac atccccgact acttcaagct
     1021 gtccttcccc gagggcttca ggtgggagcg cgtgatgaac ttcgaggacg gcggcattat
     1081 tcacgttaac caggactcct ccctgcagga cggcgtattc atctacaagg tgaagctgcg
     1141 cggcaccaac ttcccccccg acggccccgt aatgcagaag aagaccatgg gctgggagca
     1201 gtggcgcgat cctcctctcc tgtcccagcg agggcctgat gccctactca ccgtggcgct
     1261 ccggaagccc ccaggtcagc gtacggatga agaactggac ctgatctttg aagagctgct
     1321 gcatatcaag gcggtggcac acctttctaa ctcggtgaag cgggaactag ctgctgttct
     1381 gctctttgaa ccacacagca aggcaggaac tgtgttgttc agccaggggg acaagggtac
     1441 ctcgtggtac attatctgga agggatctgt caatgtggtg acccatggca aggggctggt
     1501 gaccacgttg cacgagggag atgactttgg acagctggct ctggtgaacg acgcacctcg
     1561 ggcagccacc atcatccttc gagaaaataa ctgtcacttt ctgcgtgtgg acaagcagga
     1621 cttcaaccgc atcatcaagg atgtggaatt taaagtggag actgttgatg agcggatgta
     1681 ccccgaggac ggcgccctga agagcgagat caagaagggg ctgaggctga aggacggcgg
     1741 ccactacgcc gccgaggtca agaccaccta caaggccaag aagcccgtgc agctgcccgg
     1801 cgcctacatc gtcgacatca agttggacat cgtgtcccac aacgaggact acaccatcgt
     1861 ggaacagtgc gaacgcgccg agggccgcca ctccaccggc ggcatggacg agctgtacaa
     1921 gtaaaaagct taagtttaaa ccgctgatca gcctcgactg tgccttctag ttgccagcca
     1981 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
     2041 ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg
     2101 gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct
     2161 ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg
     2221 tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     2281 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     2341 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     2401 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     2461 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     2521 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     2581 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     2641 aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
     2701 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
     2761 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     2821 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     2881 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg
     2941 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
     3001 agctcccggg agcttgtata tccattttcg gatctgatca agagacagga tgaggatcgt
     3061 ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc
     3121 tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc
     3181 tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg
     3241 aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag
     3301 ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg
     3361 ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg
     3421 caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac
     3481 atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg
     3541 acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc
     3601 ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg
     3661 aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc
     3721 aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc
     3781 gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc
     3841 ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc
     3901 caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg
     3961 aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt
     4021 cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     4081 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     4141 catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc
     4201 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     4261 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     4321 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     4381 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     4441 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     4501 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     4561 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     4621 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     4681 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     4741 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     4801 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     4861 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     4921 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     4981 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     5041 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     5101 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     5161 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     5221 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     5281 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     5341 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     5401 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     5461 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     5521 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     5581 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     5641 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     5701 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     5761 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     5821 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     5881 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     5941 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     6001 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     6061 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     6121 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     6181 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     6241 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     6301 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
     6361 cgacggatcg ggagatctcc cgatccccta tggtgcactc tcagtacaat ctgctctgat
     6421 gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc tgagtagtgc
     6481 gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat gaagaatctg
     6541 cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac gcgtt
//
