LOCUS       sequence-201361-        6674 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 6674)
  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 6674)
  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-Cdc42-T17N
FEATURES             Location/Qualifiers
     source          1..6674
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(68..87)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        259..638
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        639..842
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     793..813
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     887..906
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        887..905
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      919..928
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(925..927,928..930,931..1641)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(925..927,928..930,931..1641)
                     /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(970..991)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1231..1250)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1578..1599
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             join(1648..1695,1696..1698,1699..2223)
                     /codon_start=1
                     /product="dominant negative mutant of human Rho family 
                     small GTPase Cdc42"
                     /label=Cdc42 (T17N)
                     /translation="MQTIKCVVVGDGAVGKNCLLISYTTNKFPSEYVPTVFDNYAVTVM
                     IGGEPYTLGLFDTAGQEDYDRLRPLSYPQTDVFLVCFSVVSPSSFENVKEKWVPEITHH
                     CPKTPFLLVGTQIDLRDDPSTIEKLAKNKQKPITPETAEKLARDLKAVKYVECSALTQK
                     GLKNVFDEAILAALEPPEPKKSRRCVLL"
     promoter        complement(2251..2269)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2252..2269)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2289..2306)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2295..2519
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2565..2993
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2652..2671)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2862..2883
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3002..3022)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3007..3336
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3187..3322
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3249..3268
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3403..4197
                     /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(3457..3476)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4067..4086
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4371..4492
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4408..4427)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4462..4481
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4541..4557)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4541..4557)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4554..4576)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4565..4581
                     /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(4589..4595,4596..4613,4614..4619))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4634..4655
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4772..4789)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4943..5531)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5023..5042)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5702..6493,6494..6562))
                     /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     6325..6344
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6563..6667)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ccccgaaaag tgccacctga cgtcgacgga tcgggagatc tcccgatccc ctatggtgca
       61 ctctcagtac aatctgctct gatgccgcat agttaagcca gtatctgctc cctgcttgtg
      121 tgttggaggt cgctgagtag tgcgcgagca aaatttaagc tacaacaagg caaggcttga
      181 ccgacaattg catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac
      241 gggccagata tacgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg
      301 ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc
      361 cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca
      421 tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg
      481 cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg
      541 acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt
      601 ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca
      661 tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg
      721 tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact
      781 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag
      841 ctctctggct aactagagaa cccactgctt actggcttat cgaaattaat acgactcact
      901 atagggagac ccaagcttgc caccatggtg agcaagggcg aggagctgtt caccggggtg
      961 gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc
     1021 gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc
     1081 aagctgcccg tgccctggcc caccctcgtg accaccctga cctacggcgt gcagtgcttc
     1141 agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc
     1201 tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag
     1261 gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag
     1321 gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat
     1381 atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc
     1441 gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc
     1501 cccgtgctgc tgcccgacaa ccactacctg agcacccagt ccgccctgag caaagacccc
     1561 aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc
     1621 ggcatggacg agctgtacaa ggaattcatg cagacaatta agtgtgttgt tgtgggcgat
     1681 ggtgctgttg gtaaaaattg tctcctgata tcctacacaa caaacaaatt tccatcggaa
     1741 tatgtaccga ctgtttttga caactatgca gtcacagtta tgattggtgg agagccatat
     1801 actcttggac tttttgatac tgcagggcaa gaggattatg acagattacg accgctgagt
     1861 tatccacaaa cagatgtatt tctagtctgt ttttcagtgg tctctccatc ttcatttgaa
     1921 aacgtgaaag aaaagtgggt gcctgagata actcaccact gtccaaagac tcctttcttg
     1981 cttgttggga ctcaaattga tctcagagat gacccctcta ctattgagaa acttgccaag
     2041 aacaaacaga agcctatcac tccagagact gctgaaaagc tggcccgtga cctgaaggct
     2101 gtcaagtatg tggagtgttc tgcacttaca cagaaaggcc taaagaatgt atttgacgaa
     2161 gcaatattgg ctgccctgga gcctccagaa ccgaagaaga gccgcaggtg tgtgctgcta
     2221 tgactcgagc atgcatctag agggccctat tctatagtgt cacctaaatg ctagagctcg
     2281 ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     2341 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     2401 tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag
     2461 caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc
     2521 ttctgaggcg gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg
     2581 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     2641 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     2701 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     2761 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     2821 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     2881 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     2941 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attaattctg
     3001 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     3061 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     3121 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     3181 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     3241 atttttttta tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag
     3301 tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc
     3361 attttcggat ctgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga
     3421 ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa
     3481 cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt
     3541 ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg
     3601 ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa
     3661 gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac
     3721 cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt
     3781 gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact
     3841 cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg
     3901 ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg
     3961 acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc
     4021 atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt
     4081 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc
     4141 gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg
     4201 ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg
     4261 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     4321 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta
     4381 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     4441 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     4501 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     4561 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     4621 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     4681 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     4741 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     4801 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     4861 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     4921 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     4981 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     5041 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     5101 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     5161 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     5221 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     5281 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     5341 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     5401 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     5461 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     5521 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     5581 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     5641 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     5701 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     5761 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     5821 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     5881 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     5941 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     6001 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     6061 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     6121 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     6181 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     6241 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     6301 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     6361 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     6421 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     6481 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     6541 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     6601 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     6661 ttccgcgcac attt
//
