LOCUS       sequence-364870-        6951 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expression of a fluorescent sensor for melanin-concentrating 
            hormone in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6951)
  AUTHORS   Ino D, Tanaka Y, Hibino H, Nishiyama M
  TITLE     A fluorescent sensor for real-time measurement of extracellular 
            oxytocin dynamics in the brain.
  JOURNAL   Nat Methods. 2022 Sep 22. pii: 10.1038/s41592-022-01597-x. doi: 
            10.1038/s41592-022-01597-x.
  PUBMED    36138174
REFERENCE   2  (bases 1 to 6951)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Methods. 2022 Sep 22. pii: 10.1038/s41592-022-01597-x. doi: 
            10.1038/s41592-022-01597-x."
COMMENT     Sequence Label: pCIS.MTRIA-MCH
FEATURES             Location/Qualifiers
     source          1..6951
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        306..581
                     /label=chicken beta-actin promoter
     intron          582..1598
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1522..1544
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1606..1625
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     1654..1670
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     1655..1671
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2630..2651
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2757..2778)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3478..3494)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3478..3494)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(3583..3602)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3648..3703
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3702..3721)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     promoter        complement(4058..4076)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4097..4113)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4097..4113)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4110..4132)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4121..4137
                     /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(join(4145..4151,4152..4169,4170..4175))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4190..4211
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4328..4345)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4499..5087)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4579..4598)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5258..6049,6050..6118))
                     /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     5881..5900
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6119..6223)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6250..6705
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6337..6356)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6547..6568
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6831..6853
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6845..6862
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6846..6862
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6869..6888
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6869..6887
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtcgag gtgagcccca cgttctgctt cactctcccc atctcccccc cctccccacc
      361 cccaattttg tatttattta ttttttaatt attttgtgca gcgatggggg cggggggggg
      421 gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg gggcggggcg aggcggagag
      481 gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt tccttttatg gcgaggcggc
      541 ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc gggagtcgct gcgcgctgcc
      601 ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc gccccggctc tgactgaccg
      661 cgttactccc acaggtgagc gggcgggacg gcccttctcc tccgggctgt aattagcgct
      721 tggtttaatg acggcttgtt tcttttctgt ggctgcgtga aagccttgag gggctccggg
      781 agggcccttt gtgcgggggg agcggctcgg ggggtgcgtg cgtgtgtgtg tgcgtgggga
      841 gcgccgcgtg cggctccgcg ctgcccggcg gctgtgagcg ctgcgggcgc ggcgcggggc
      901 tttgtgcgct ccgcagtgtg cgcgagggga gcgcggccgg gggcggtgcc ccgcggtgcg
      961 gggggggctg cgaggggaac aaaggctgcg tgcggggtgt gtgcgtgggg gggtgagcag
     1021 ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca cccccctccc cgagttgctg
     1081 agcacggccc ggcttcgggt gcggggctcc gtacggggcg tggcgcgggg ctcgccgtgc
     1141 cgggcggggg gtggcggcag gtgggggtgc cgggcggggc ggggccgcct cgggccgggg
     1201 agggctcggg ggaggggcgc ggcggccccc ggagcgccgg cggctgtcga ggcgcggcga
     1261 gccgcagcca ttgcctttta tggtaatcgt gcgagagggc gcagggactt cctttgtccc
     1321 aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac cccctctagc gggcgcgggg
     1381 cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga gggccttcgt gcgtcgccgc
     1441 gccgccgtcc ccttctccct ctccagcctc ggggctgtcc gcggggggac ggctgccttc
     1501 gggggggacg gggcagggcg gggttcggct tctggcgtgt gaccggcggc tctagagcct
     1561 ctgctaacca tgttcatgcc ttcttctttt tcctacagct cctgggcaac gtgctggtta
     1621 ttgtgctgtc tcatcatttt ggcaaaggat tcctcgaggt cgacggtatc gataagcttg
     1681 atatcgaatt caccatggat ctgcaagcct cgttgctgtc cactggcccc aatgccagca
     1741 acatctccga tggccaggat aatttcacat tggcggggcc acctcctcgc acaaggagtg
     1801 tctcctacat caacatcatc atgccttcag tgtttggtac catctgtctc ctgggcattg
     1861 tgggaaactc cacagtcatt tttgccgtgg tgaagaaatc caagctgcac tggtgcagca
     1921 acgtccctga catcttcatc atcaacctct ctgtggtgga tctgcttttc ctgctgggca
     1981 tgcctttcat gatccaccag ctcatgggta atggtgtctg gcactttggg gaaaccatgt
     2041 gcaccctcat cacagccatg gacgccaaca gtcagttcac cagcacctac atcctgactg
     2101 ctatggccat tgaccgctac ttggccaccg tccatcccat ctcctccacc aagttccgga
     2161 agccctccat ggccaccctg gtgatctgcc tcctgtgggc tctctcgttc attagcatca
     2221 ctcctgtgtg gctctatgcc aggcttatcc ccttcccagg gggtgctgtg ggctgtggca
     2281 tccgcctacc aaacccagat actgatcttt actggttcac tctgtatcag tttttcctgg
     2341 ccttcgccct tccgtttgtg gtcatcactg ctgcgtacgt gaaaatatgc tacaaaatat
     2401 ggcgaaattt caaagggaaa ttaaacgtct atatcaaggc cgacaagcag aagaacggca
     2461 tcaaggcgaa cttccacatc cgccacaaca tcgaggacgg cggcgtgcag ctcgcctacc
     2521 actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc
     2581 tgagcgtgca gaccaaactt tcgaaagacc ccaacgagaa gcgcgatcac atggtcctgc
     2641 tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagggcggta
     2701 ccggagggat gatggtgaga aagggcgagg agctgttcac cggggtggtg cccatcctgg
     2761 tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag ggtgagggcg
     2821 atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc
     2881 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg
     2941 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac atccaggagc
     3001 gcaccatctt cttcaaggac gacggccact acaagacccg cgccgaggtg aagttcgagg
     3061 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
     3121 tcctggggca caagctggag tacaactcag taatctccaa ggcaaagata cgcacagtga
     3181 aaatggccat tgccatctgt ctggtcttct ttgtgtgctg ggcgccctac tacgtgctgc
     3241 agctgaccca gttgtccatc agccgcccga ccctcacatt cgtctacctg tacaatgcgg
     3301 ctatcagctt gggctatgcc aacagctgcc tcaatccctt tgtgtacata gtactctgtg
     3361 agacctttcg aaaacgcttg gtgctgtcgg tgaagcccgc ggcccagggg cagcttcgca
     3421 cggtcagcaa tgctcagaca gctgacgagg agaggacaga aagcaaaggc acctgaggat
     3481 ccactagttc tagagcggcc gcttcactcc tcaggtgcag gctgcctatc agaaggtggt
     3541 ggctggtgtg gccaatgccc tggctcacaa ataccactga gatctttttc cctctgccaa
     3601 aaattatggg gacatcatga agccccttga gcatctgact tctggctaat aaaggaaatt
     3661 tattttcatt gcaatagtgt gttggaattt tttgtgtctc tcactcggaa ggacatatgg
     3721 gagggcaaat catttaaaac atcagaatga gtatttggtt tagagtttgg caacatatgc
     3781 ccatatgctg gctgccatga acaaaggttg gctataaaga ggtcatcagt atatgaaaca
     3841 gccccctgct gtccattcct tattccatag aaaagccttg acttgaggtt agattttttt
     3901 tatattttgt tttgtgttat ttttttcttt aacatcccta aaattttcct tacatgtttt
     3961 actagccaga tttttcctcc tctcctgact actcccagtc atagctgtcc ctcttctctt
     4021 atgaagatcc ctcgacctgg agctccagct tttgttccct ttagtgaggg ttaatttcga
     4081 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
     4141 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
     4201 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     4261 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     4321 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     4381 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     4441 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     4501 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     4561 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     4621 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     4681 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     4741 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     4801 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     4861 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     4921 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     4981 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     5041 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     5101 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     5161 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     5221 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     5281 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     5341 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     5401 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     5461 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     5521 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     5581 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     5641 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     5701 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     5761 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     5821 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     5881 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     5941 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     6001 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     6061 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     6121 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     6181 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     6241 tgccacctga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     6301 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     6361 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     6421 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     6481 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     6541 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt
     6601 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     6661 aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttccatt cgccattcag
     6721 gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct tcgctattac gccagctggc
     6781 gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt cccagtcacg
     6841 acgttgtaaa acgacggcca gtgaattgta atacgactca ctatagggcg aattgggtac
     6901 ctagtaatca attacggggt cattagttca tagcccatat atggagttcc g
//
