LOCUS       sequence-220047-        7837 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7837)
  AUTHORS   Pertz O, Hodgson L, Klemke RL, Hahn KM
  TITLE     Spatiotemporal dynamics of RhoA activity in migrating cells.
  JOURNAL   Nature. 2006 Apr 20. 440(7087):1069-72.
  PUBMED    16547516
REFERENCE   2  (bases 1 to 7837)
  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: "Nature. 
            2006 Apr 20. 440(7087):1069-72."
COMMENT     Sequence Label: pBabe-Sin-Puro-Tet-RhoA FLARE.sc Biosensor
FEATURES             Location/Qualifiers
     source          1..7837
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    534..733
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             934..1350
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1280..1302
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1318..1334
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     protein_bind    join(1391..1409,1410..1432,1433..1451,1452..1474,1475..1493,
                     1494..1516,1517..1535,1536..1558,1559..1577,1578..1600,
                     1601..1619,1620..1642,1643..1661)
                     /label=tetracycline response element
                     /bound_moiety="tetracycline repressor TetR"
                     /note="contains seven copies of the tetracycline operator 
                     tetO"
     promoter        1694..1732
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1729..1753
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             1851..1868
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             join(2133..2135,2136..2138,2139..2849)
                     /codon_start=1
                     /product="cyan variant of GFP"
                     /label=CFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     AHFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2922..2924,2925..2927,2928..3638)
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             3645..4226
                     /codon_start=1
                     /product="human Rho family small GTPase RhoA"
                     /label=RhoA
                     /translation="MAAIRKKLVIVGDGACGKTCLLIVFSKDQFPEVYVPTVFENYVAD
                     IEVDGKQVELALWDTAGQEDYDRLRPLSYPDTDVILMCFSIDSPDSLENIPEKWTPEVK
                     HFCPNVPIILVGNKKDLRNDEHTRRELAKMKQEPVKPEEGRDMANRIGAFGYMECSAKT
                     KDGVREVFEMATRAALQARRGKKKSGCLVL"
     protein_bind    4243..4259
                     /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        4292..4791
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(4327..4350)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     4724..4743
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             4814..5413
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4814..4833)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5310..5330
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(5922..5939)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6093..6681)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6173..6192)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6841..7632,7633..7701))
                     /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     7464..7483
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7702..7806)
                     /gene="bla"
                     /label=AmpR promoter
     LTR             join(7837,1..469)
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
ORIGIN
        1 tgaatatggg ccaaacagga tatctgtggt aagcagttcc tgccccggct cagggccaag
       61 aacagatgga acagctgaat atgggccaaa caggatatct gtggtaagca gttcctgccc
      121 cggctcaggg ccaagaacag atggtcccca gatgcggtcc agccctcagc agtttctaga
      181 gaaccatcag atgtttccag ggtgccccaa ggacctgaaa tgaccctgtg ccttatttga
      241 actaaccaat cagttcgctt ctcgcttctg ttcgcgcgct tctgctcccc gagctcaata
      301 aaagagccca caacccctca ctcggggcgc cagtcctccg attgactgag tcgcccgggt
      361 acccgtgtat ccaataaacc ctcttgcagt tgcatccgac ttgtggtctc gctgttcctt
      421 gggagggtct cctctgagtg attgactacc cgtcagcggg ggtctttcat ttgggggctc
      481 gtccgggatc gggagacccc tgcccaggga ccaccgaccc accaccggga ggcaagctgg
      541 ccagcaactt atctgtgtct gtccgattgt ctagtgtcta tgactgattt tatgcgcctg
      601 cgtcggtact agttagctaa ctagctctgt atctggcgga cccgtggtgg aactgacgag
      661 ttctgaacac ccggccgcaa ccctgggaga cgtcccaggg actttggggg ccgtttttgt
      721 ggcccgacct gaggaaggga gtcgatgtgg aatccgaccc cgtcaggata tgtggttctg
      781 gtaggagacg agaacctaaa acagttcccg cctccgtctg aatttttgct ttcggtttgg
      841 aaccgaagcc gcgcgtcttg tctgctgcag cgctgcagca tcgttctgtg ttgtctctgt
      901 ctgactgtgt ttctgtattt gtctgaaaat tagggccaga ctgttaccac tcccttaagt
      961 ttgaccttag gtcactggaa agatgtcgag cggatcgctc acaaccagtc ggtagatgtc
     1021 aagaagagac gttgggttac cttctgctct gcagaatggc caacctttaa cgtcggatgg
     1081 ccgcgagacg gcacctttaa ccgagacctc atcacccagg ttaagatcaa ggtcttttca
     1141 cctggcccgc atggacaccc agaccaggtc ccctacatcg tgacctggga agccttggct
     1201 tttgaccccc ctccctgggt caagcccttt gtacacccta agcctccgcc tcctcttcct
     1261 ccatccgccc cgtctctccc ccttgaacct cctcgttcga ccccgcctcg atcctccctt
     1321 tatccagccc tcactccttc tctaggcgcc ggccggatcc acgaggccct ttcgtcgtcg
     1381 agtttaccac tccctatcag tgatagagaa aagtgaaagt cgagtttacc actccctatc
     1441 agtgatagag aaaagtgaaa gtcgagttta ccactcccta tcagtgatag agaaaagtga
     1501 aagtcgagtt taccactccc tatcagtgat agagaaaagt gaaagtcgag tttaccactc
     1561 cctatcagtg atagagaaaa gtgaaagtcg agtttaccac tccctatcag tgatagagaa
     1621 aagtgaaagt cgagtttacc actccctatc agtgatagag aaaagtgaaa gtcgagctcg
     1681 gtacccgggt cgaggtaggc gtgtacggtg ggaggcctat ataagcagag ctcgtttagt
     1741 gaaccgtcag atcgcctgga gacgccatcc acgctgtttt gacctccata gaagacaccg
     1801 ggaccgatcc agcctccgcg gccccagatc cgtaaagtga attcatggca caccatcacc
     1861 accatcacgg tagtggcatc ctggaggacc tcaatatgct ctacatccgg cagatggcac
     1921 tcagcctgga ggacacagag ctgcagagga aactagatca tgagatccgg atgagggatg
     1981 gggcctgcaa gctgctggca gcctgctccc agcgagagca ggctctggaa gccaccaaga
     2041 gcctgctggt gtgcaacagc cgtattctca gctacatggg tgagctgcag cggcgaaagg
     2101 aggcccaggt gctggagaag acaggcgggg gaatggtgag caagggcgag gagctgttca
     2161 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aggttcagcg
     2221 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca
     2281 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc tggggcgtgc
     2341 agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
     2401 ccgaaggcta cgtccaggag cgtaccatct tcttcaagga cgacggcaac tacaagaccc
     2461 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
     2521 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac atcagccaca
     2581 acgtctatat caccgccgac aagcagaaga acggcatcaa ggcccacttc aagatccgcc
     2641 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     2701 gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca
     2761 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     2821 tcactctcgg catggacgag ctgtacaagg gatctacttc tggttctggt aaacctggtt
     2881 ctggtgaagg ttctactaaa ggtggtagtg gcgcggccgc tatggtgagc aagggcgagg
     2941 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca
     3001 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt
     3061 tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accttcggct
     3121 acggcctgat gtgcttcgcc cgctaccccg accacatgaa gcagcacgac ttcttcaagt
     3181 ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact
     3241 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga
     3301 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca
     3361 acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca
     3421 agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca
     3481 cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc taccagtccg
     3541 ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg
     3601 ccgccgggat cactctcggc atggacgagc tgtacaaggg gggaatggct gccatccgga
     3661 agaaactggt gattgttggt gatggagcct gtggaaagac atgcttgctc atagtcttca
     3721 gcaaggacca gttcccagag gtgtatgtgc ccacagtgtt tgagaactat gtggcagata
     3781 tcgaggtgga tggaaagcag gtagagttgg ctttgtggga cacagctggg caggaagatt
     3841 atgatcgcct gaggcccctc tcctacccag ataccgatgt tatactgatg tgtttttcca
     3901 tcgacagccc tgatagttta gaaaacatcc cagaaaagtg gaccccagaa gtcaagcatt
     3961 tctgtcccaa cgtgcccatc atcctggttg ggaataagaa ggatcttcgg aatgatgagc
     4021 acacaaggcg ggagctagcc aagatgaagc aggagccggt gaaacctgaa gaaggcagag
     4081 atatggcaaa caggattggc gcttttgggt acatggagtg ttcagcaaag accaaagatg
     4141 gagtgagaga ggtttttgaa atggctacga gagctgctct gcaagctaga cgtgggaaga
     4201 aaaaatctgg ttgccttgtc ttgtgataac tcgagtgtgg aattgtgagc ggataacaat
     4261 ttgtcgaggg cccctgcagg tcaattctac cgggtagggg aggcgctttt cccaaggcag
     4321 tctggagcat gcgctttagc agccccgctg ggcacttggc gctacacaag tggcctctgg
     4381 cctcgcacac attccacatc caccggtagg cgccaaccgg ctccgttctt tggtggcccc
     4441 ttcgcgccac cttctactcc tcccctagtc aggaagttcc cccccgcccc gcagctcgcg
     4501 tcgtgcagga cgtgacaaat ggaagtagca cgtctcacta gtctcgtgca gatggacagc
     4561 accgctgagc aatggaagcg ggtaggcctt tggggcagcg gccaatagca gctttgctcc
     4621 ttcgctttct gggctcagag gctgggaagg ggtgggtccg ggggcgggct caggggcggg
     4681 ctcaggggcg gggcgggcgc ccgaaggtcc tccggaggcc cggcattctg cacgcttcaa
     4741 aagcgcacgt ctgccgcgct gttctcctct tcctcatctc cgggcctttc gacctgcagc
     4801 caataagctt accatgaccg agtacaagcc cacggtgcgc ctcgccaccc gcgacgacgt
     4861 ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac taccccgcca cgcgccacac
     4921 cgtcgatccg gaccgccaca tcgagcgggt caccgagctg caagaactct tcctcacgcg
     4981 cgtcgggctc gacatcggca aggtgtgggt cgcggacgac ggcgccgcgg tggcggtctg
     5041 gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc cgcgcatggc
     5101 cgagttgagc ggttcccggc tggccgcgca gcaacagatg gaaggcctcc tggcgccgca
     5161 ccggcccaag gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg accaccaggg
     5221 caagggtctg ggcagcgccg tcgtgctccc cggagtggag gcggccgagc gcgccggggt
     5281 gcccgccttc ctggagacct ccgcgccccg caacctcccc ttctacgagc ggctcggctt
     5341 caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc acctggtgca tgacccgcaa
     5401 gcccggtgcc tgacgcccgc cccacgaccc gcagcgcccg accgaaagga gcgcacgacc
     5461 ccatgcatcg ataaaataaa agattttatt tagtctccag aaaaaggggg gaatgaaaga
     5521 ccccacctgt aggtttggca agctagctta agtaacgcca ttttgcaagg catggaaaaa
     5581 tacataactg agaatagaga agttcagatc aaggtaggag atccctgagc ccacaacccc
     5641 tcactcgggg cgccagtcct ccgattgact gagtcgcccg ggtacccgtg tatccaataa
     5701 accctcttgc agttgcatcc gacttgtggt ctcgctgttc cttgggaggg tctcctctga
     5761 gtgattgact acccgtcagc gggggtcttt cacatgcagc atgtatcaaa attaatttgg
     5821 ttttttttct taagtattta cattaaatgg ccatagttgc attaatgaat cggccaacgc
     5881 gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg
     5941 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     6001 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     6061 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     6121 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     6181 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     6241 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     6301 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     6361 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     6421 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     6481 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta
     6541 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     6601 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     6661 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     6721 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     6781 tagatccttt tgcggccgca aatcaatcta aagtatatat gagtaaactt ggtctgacag
     6841 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     6901 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     6961 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     7021 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     7081 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     7141 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     7201 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     7261 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     7321 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     7381 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     7441 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     7501 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     7561 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     7621 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     7681 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     7741 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     7801 tccgcgcaca tttccccgaa aagtgccacc tgcagcc
//
