LOCUS       sequence-170000-        8229 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  MESA target chain with V2-MESA ectodomain, 35 extracellular linkers,
            a flag tag, M cleavage sequence, and tTA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8229)
  AUTHORS   Schwarz KA, Daringer NM, Dolberg TB, Leonard JN
  TITLE     Rewiring human cellular input-output using modular extracellular 
            sensors.
  JOURNAL   Nat Chem Biol. 2016 Dec 12. doi: 10.1038/nchembio.2253.
  PUBMED    27941759
REFERENCE   2  (bases 1 to 8229)
  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 Chem 
            Biol. 2016 Dec 12. doi: 10.1038/nchembio.2253."
COMMENT     Sequence Label: V2-MESA-35F-M-tTA
FEATURES             Location/Qualifiers
     source          1..8229
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        175..554
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        555..758
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     709..729
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     803..822
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        803..821
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1780..1800
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQM"
     CDS             1798..2421
                     /codon_start=1
                     /gene="tetR"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     CDS             join(1801..2421,2422..2805)
                     /codon_start=1
                     /product="tetracycline-controlled transactivator, 
                     comprising a fusion of the tetracycline repressor TetR with
                     the C-terminal activation domain of herpes simplex virus 
                     VP16"
                     /label=tTA
                     /note="In the Tet-Off(R) system, tTA binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the absence of tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGSAYSRARTKNNYGSTI
                     EGLLDLPDDDAPEEAGLAAPRLSFLPAGHTRRLSTAPPTDVSLGDELHLDGEDVAMAHA
                     DALDDFDLDMLGDGDSPGPGFTPHDSAPYGALDMADFEFEQMFTDALGIDEYGG"
     primer_bind     complement(1874..1893)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     CDS             join(2836..2838,2839..2841,2842..3555)
                     /codon_start=1
                     /product="enhanced blue variant of GFP (Ai et al., 2007)"
                     /label=EBFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLSHGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGVDFKEDGNILGHKLEYNFNSHNIYIMAVKQKNGIK
                     VNFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDSHYLSTQSVLSKDPNEKRDHMVLL
                     EFRTAAGITLGMDELYK"
     primer_bind     complement(2881..2902)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     polyA_signal    3676..4068
                     /label=beta-globin poly(A)
                     /note="human beta-globin polyadenylation signal"
     rep_origin      4976..5489
                     /direction=RIGHT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5063..5082)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5273..5294
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5970..5989)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6089..6111
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6149..6167)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6235..6339
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6340..6408,6409..7200)
                     /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     complement(6558..6577)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7371..7959
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7860..7879
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   join(8199..8229,1..99)
                     /label=AAV2 ITR
ORIGIN
        1 gggcaaagcc cgggcgtcgg gcgacctttg gtcgcccggc ctcagtgagc gagcgagcgc
       61 gcagagaggg agtggccaac tccatcacta ggggttcctt gtagttaatg attaacccgc
      121 catgctactt atctacgtag ccatgctcta gggatccggc cagatatacg cgttgacatt
      181 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      241 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      301 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      361 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      421 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      481 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      541 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      601 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      661 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      721 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      781 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      841 gccatcatgg cttggaccag cttaattctt agcttgcttg ccctgtgcag cggtgcatca
      901 tctgacatac agatgactca gtcacctagc agtctgtctg cttcagtcgg tgatagggta
      961 actatcacct gtcgggcatc tcaggtgatc agaaggtctc tggcctggta tcagcagaag
     1021 cccggcaagg ctcctaagtt gcttatctat gcagcttcaa acttggccag tggggtgcct
     1081 agccggttta gcggaagtgg ttctggcaca gacttcactt tgacaatcag ttccctccaa
     1141 ccggaggatt tcgctaccta ctactgtcag cagtccaaca ccagccctct gacgtttgga
     1201 cagggcacta aagtggagat taagagggga ggcggaggct ccggtggggg aggctctggt
     1261 ggtggaggta gcgaggttca gctggtggaa agtggtggcg gactagtaca gccaggcggc
     1321 tccctgcggc tgagttgcgc cgcgagtggc ttttctataa acggatcctg gatcttctgg
     1381 gtcaggcaag cccctgggaa aggattggag tgggttggag ccatttggcc tttcggtggg
     1441 tatactcact atgcagatag cgtgaaaggg aggttcacca tctctgcaga cacaagcaag
     1501 aacacggcct acctacagat gaactctctg agggcagagg acactgccgt ctactattgc
     1561 gcccgatggg gacactccac aagcccttgg gccatggact attggggaca ggggacattg
     1621 gtaggcgccg gatctggagg agactacaag gacgatgacg acaacggcgg atcggggggg
     1681 tctggtggta gtggcggttc ggggggaggt accggtctgg tcgtggttgc tggagtcctg
     1741 ttttgttatg gcttgctagt gacagtggct ctttgtgttg aaaacctgta ttttcagatg
     1801 atgtctagat tagataaaag taaagtgatt aacagcgcat tagagctgct taatgaggtc
     1861 ggaatcgaag gtttaacaac ccgtaaactc gcccagaagc taggtgtaga gcagcctaca
     1921 ttgtattggc atgtaaaaaa taagcgggct ttgctcgacg ccttagccat tgagatgtta
     1981 gataggcacc atactcactt ttgcccttta gaaggggaaa gctggcaaga ttttttacgt
     2041 aataacgcta aaagttttag atgtgcttta ctaagtcatc gcgatggagc aaaagtacat
     2101 ttaggtacac ggcctacaga aaaacagtat gaaactctcg aaaatcaatt agccttttta
     2161 tgccaacaag gtttttcact agagaatgca ttatatgcac tcagcgctgt ggggcatttt
     2221 actttaggtt gcgtattgga agatcaagag catcaagtcg ctaaagaaga aagggaaaca
     2281 cctactactg atagtatgcc gccattatta cgacaagcta tcgaattatt tgatcaccaa
     2341 ggtgcagagc cagccttctt attcggcctt gaattgatca tatgcggatt agaaaaacaa
     2401 cttaaatgtg aaagtgggtc cgcgtacagc cgcgcgcgta cgaaaaacaa ttacgggtct
     2461 accatcgagg gcctgctcga tctcccggac gacgacgccc ccgaagaggc ggggctggcg
     2521 gctccgcgcc tgtcctttct ccccgcggga cacacgcgca gactgtcgac ggcccccccg
     2581 accgatgtca gcctggggga cgagctccac ttagacggcg aggacgtggc gatggcgcat
     2641 gccgacgcgc tagacgattt cgatctggac atgttggggg acggggattc cccgggtccg
     2701 ggatttaccc cccacgactc cgccccctac ggcgctctgg atatggccga cttcgagttt
     2761 gagcagatgt ttaccgatgc ccttggaatt gacgagtacg gtgggggtac cggcggaggc
     2821 tccggtggtg gctctatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc
     2881 ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgagggg cgagggcgag
     2941 ggcgatgcca ccaacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc
     3001 gtgccctggc ccaccctcgt gaccaccctg agccacggcg tgcagtgctt cgcccgctac
     3061 cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag
     3121 gagcgcacca tcttcttcaa ggacgacggc acctacaaga cccgcgccga ggtgaagttc
     3181 gagggcgaca ccctagtgaa ccgcatcgag ctgaagggcg tcgacttcaa ggaggacggc
     3241 aacatcctgg ggcacaagct ggagtacaac ttcaacagcc acaacatcta tatcatggcc
     3301 gtcaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacgt ggaggacggc
     3361 agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg
     3421 ctgcccgaca gccactacct gagcacccag tccgtgctga gcaaagaccc caacgagaag
     3481 cgcgatcaca tggtcctgct ggagttccgc accgccgccg ggatcactct cggcatggac
     3541 gagctgtaca agtaatagcg gccgccaccg cggtggagct gtttaaacct cgagcaattc
     3601 accccaccag tgcaggctgc ctatcagaaa gtggtggctg gtgtggctaa tgccctggcc
     3661 cacaagtatc actaagctcg ctttcttgct gtccaatttc tattaaaggt tcctttgttc
     3721 cctaagtcca actactaaac tggggatatt atgaagggcc ttgagcatct ggattctgcc
     3781 taataaaaaa catttatttt cattgcaatg atgtatttaa attatttctg aatattttac
     3841 taaaaaggga atgtgggagg tcagtgcatt taaaacataa agaaatgaag agctagttca
     3901 aaccttggga aaatacacta tatcttaaac tccatgaaag aaggtgaggc tgcaaacagt
     3961 aatgcacatt ggcaacagcc cctgatgcct atgccttatt catccctcag aaaaggattc
     4021 aagtagaggc ttgatttgga ggttaaagtt ttgctatgct gtattttaca ttacttattg
     4081 ttttagctgt cctcatgaat gtcttttcac tacccatttg cttatcctgc atctctcagc
     4141 cttgactcca ctcagttctc ttgcttagag ataccacctt tcccctgaag tgttccttcc
     4201 atgttttacg gcgagatggt ttctcctcgc ctggccactc agccttagtt gtctctgttg
     4261 tcttatagag gtctacttga agaaggaaaa acagggggca tggtttgact gtcctgtgag
     4321 cccttcttcc ctgcctcccc cactcacagt gacccggaat ccctcgacat ggcagtctag
     4381 agcatggcta cgtagataag tagcatggcg ggttaatcat taactacaag gaacccctag
     4441 tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa
     4501 aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga gcgcgccagc
     4561 tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat
     4621 ggcgaatgga attccagacg attgagcgtc aaaatgtagg tatttccatg agcgtttttc
     4681 ctgttgcaat ggctggcggt aatattgttc tggatattac cagcaaggcc gatagtttga
     4741 gttcttctac tcaggcaagt gatgttatta ctaatcaaag aagtattgcg acaacggtta
     4801 atttgcgtga tggacagact cttttactcg gtggcctcac tgattataaa aacacttctc
     4861 aggattctgg cgtaccgttc ctgtctaaaa tccctttaat cggcctcctg tttagctccc
     4921 gctctgattc taacgaggaa agcacgttat acgtgctcgt caaagcaacc atagtacgcg
     4981 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     5041 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     5101 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     5161 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     5221 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     5281 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     5341 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     5401 aattttaaca aaatattaac gtttacaatt taaatatttg cttatacaat cttcctgttt
     5461 ttggggcttt tctgattatc aaccggggta catatgattg acatgctagt tttacgatta
     5521 ccgttcatcg attctcttgt ttgctccaga ctctcaggca atgacctgat agcctttgta
     5581 gagacctctc aaaaatagct accctctccg gcatgaattt atcagctaga acggttgaat
     5641 atcatattga tggtgatttg actgtctccg gcctttctca cccgtttgaa tctttaccta
     5701 cacattactc aggcattgca tttaaaatat atgagggttc taaaaatttt tatccttgcg
     5761 ttgaaataaa ggcttctccc gcaaaagtat tacagggtca taatgttttt ggtacaaccg
     5821 atttagcttt atgctctgag gctttattgc ttaattttgc taattctttg ccttgcctgt
     5881 atgatttatt ggatgttgga attcctgatg cggtattttc tccttacgca tctgtgcggt
     5941 atttcacacc gcatatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
     6001 cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca
     6061 tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg
     6121 tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt ataggttaat
     6181 gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga
     6241 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     6301 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     6361 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     6421 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     6481 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     6541 agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     6601 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     6661 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     6721 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     6781 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     6841 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     6901 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     6961 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     7021 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     7081 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     7141 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     7201 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     7261 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     7321 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     7381 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     7441 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     7501 cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct
     7561 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     7621 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     7681 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     7741 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     7801 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     7861 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     7921 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     7981 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     8041 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     8101 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg
     8161 cctctccccg cgcgttggcc gattcattaa tgcagcagct gcgcgctcgc tcgctcactg
     8221 aggccgccc
//
