LOCUS       sequence-430582-        6680 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 6680)
  AUTHORS   Subauste MC, Von Herrath M, Benard V, Chamberlain CE, Chuang TH, Chu
            K, Bokoch GM, Hahn KM
  TITLE     Rho family proteins modulate rapid apoptosis induced by cytotoxic T 
            lymphocytes and Fas.
  JOURNAL   J Biol Chem. 2000 Mar 31. 275(13):9725-33.
  PUBMED    10734125
REFERENCE   2  (bases 1 to 6680)
  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: "J Biol 
            Chem. 2000 Mar 31. 275(13):9725-33."
COMMENT     Sequence Label: pcDNA3-EGFP-RhoA-wt
FEATURES             Location/Qualifiers
     source          1..6680
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(247..265)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(248..265)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(285..302)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    291..515
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      561..989
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(648..667)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     858..879
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(998..1018)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1003..1332
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1183..1318
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1245..1264
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             1399..2193
                     /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(1453..1472)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2063..2082
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    2367..2488
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2404..2423)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2458..2477
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2537..2553)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2537..2553)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2550..2572)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2561..2577
                     /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(2585..2591,2592..2609,2610..2615))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2630..2651
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2768..2785)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2939..3527)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3019..3038)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3698..4489,4490..4558))
                     /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     4321..4340
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4559..4663)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4738..4757)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        4929..5308
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5309..5512
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5463..5483
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5557..5576
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5557..5575
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      5589..5598
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(5595..5597,5598..5600,5601..6311)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(5595..5597,5598..5600,5601..6311)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(5640..5661)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5901..5920)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6248..6269
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             join(6318..6680,1..219)
                     /codon_start=1
                     /product="human Rho family small GTPase RhoA"
                     /label=RhoA
                     /translation="MAAIRKKLVIVGDGACGKTCLLIVFSKDQFPEVYVPTVFENYVAD
                     IEVDGKQVELALWDTAGQEDYDRLRPLSYPDTDVILMCFSIDSPDSLENIPEKWTPEVK
                     HFCPNVPIILVGNKKDLRNDEHTRRELAKMKQEPVKPEEGRDMANRIGAFGYMECSAKT
                     KDGVREVFEMATRAALQARRGKKKSGCLVL"
ORIGIN
        1 cggaatgatg agcacacaag gcgggagcta gccaagatga agcaggagcc ggtgaaacct
       61 gaagaaggca gagatatggc aaacaggatt ggcgcttttg ggtacatgga gtgttcagca
      121 aagaccaaag atggagtgag agaggttttt gaaatggcta cgagagctgc tctgcaagct
      181 agacgtggga agaaaaaatc tggttgcctt gtcttgtgac tcgagcatgc atctagaggg
      241 ccctattcta tagtgtcacc taaatgctag agctcgctga tcagcctcga ctgtgccttc
      301 tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc
      361 cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg
      421 tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa
      481 tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg
      541 gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
      601 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
      661 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
      721 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
      781 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
      841 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
      901 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
      961 gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca gttagggtgt
     1021 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     1081 gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     1141 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg
     1201 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     1261 gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
     1321 ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga tcaagagaca
     1381 ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct
     1441 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
     1501 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
     1561 ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc cacgacgggc
     1621 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
     1681 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
     1741 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac
     1801 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
     1861 caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc
     1921 aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg
     1981 aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg
     2041 gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc
     2101 gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc
     2161 gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg
     2221 accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa
     2281 ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc
     2341 tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat
     2401 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
     2461 gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga
     2521 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
     2581 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
     2641 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     2701 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     2761 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     2821 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     2881 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     2941 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     3001 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     3061 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     3121 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     3181 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     3241 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     3301 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     3361 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     3421 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     3481 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     3541 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     3601 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     3661 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     3721 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     3781 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     3841 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     3901 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     3961 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     4021 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     4081 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     4141 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     4201 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     4261 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     4321 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     4381 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     4441 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     4501 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     4561 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     4621 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     4681 tgccacctga cgtcgacgga tcgggagatc tcccgatccc ctatggtgca ctctcagtac
     4741 aatctgctct gatgccgcat agttaagcca gtatctgctc cctgcttgtg tgttggaggt
     4801 cgctgagtag tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg
     4861 catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata
     4921 tacgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
     4981 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
     5041 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
     5101 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
     5161 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
     5221 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
     5281 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg
     5341 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
     5401 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt
     5461 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctctctggct
     5521 aactagagaa cccactgctt actggcttat cgaaattaat acgactcact atagggagac
     5581 ccaagcttgc caccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc
     5641 tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg
     5701 gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg
     5761 tgccctggcc caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc
     5821 ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg
     5881 agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg
     5941 agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca
     6001 acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg
     6061 acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca
     6121 gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc
     6181 tgcccgacaa ccactacctg agcacccagt ccgccctgag caaagacccc aacgagaagc
     6241 gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg
     6301 agctgtacaa ggaattcatg gctgccatcc ggaagaaact ggtgattgtt ggtgatggag
     6361 cctgtggaaa gacatgcttg ctcatagtct tcagcaagga ccagttccca gaggtgtatg
     6421 tgcccacagt gtttgagaac tatgtggcag atatcgaggt ggatggaaag caggtagagt
     6481 tggctttgtg ggacacagct gggcaggaag attatgatcg cctgaggccc ctctcctacc
     6541 cagataccga tgttatactg atgtgttttt ccatcgacag ccctgatagt ttagaaaaca
     6601 tcccagaaaa gtggacccca gaagtcaagc atttctgtcc caacgtgccc atcatcctgg
     6661 ttgggaataa gaaggatctt
//
