LOCUS       sequence-47142-e        6716 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 6716)
  AUTHORS   Kiyomitsu T, Cheeseman IM
  TITLE     Chromosome- and spindle-pole-derived signals generate an intrinsic 
            code for spindle position and orientation.
  JOURNAL   Nat Cell Biol. 2012 Feb 12;14(3):311-7. doi: 10.1038/ncb2440.
  PUBMED    22327364
REFERENCE   2  (bases 1 to 6716)
  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"; doi: "10"; journalName: 
            "Nat Cell Biol"; date: "2012-02-12- 12"; volume: "14"; issue: "3"; 
            pages: "311-7"
COMMENT     Sequence Label: pTK25
FEATURES             Location/Qualifiers
     source          1..6716
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             670..1377
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(815..833)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1083..1102
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1423..1443
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1456..1500
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     polyA_signal    3504..3625
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3541..3560)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3595..3614
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3632..4087)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3769..3790)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3981..4000
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4114..4218
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4220..4577
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4220..4411
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4243..4263)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4428..4563
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4612..5406
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4612..5406
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4666..4685)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5276..5295
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5594..5613)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5638..5685
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6014..6602
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6503..6522
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcggcc
      601 accatgctgt gctgtatgag aagaaccaaa caggttgaaa aaaatgatga cgaccaaaag
      661 attggcggca tggtgagcaa gggcgaggag gataacatgg ccatcatcaa ggagttcatg
      721 cgcttcaagg tgcacatgga gggctccgtg aacggccacg agttcgagat cgagggcgag
      781 ggcgagggcc gcccctacga gggcacccag accgccaagc tgaaggtgac caagggtggc
      841 cccctgccct tcgcctggga catcctgtcc cctcagttca tgtacggctc caaggcctac
      901 gtgaagcacc ccgccgacat ccccgactac ttgaagctgt ccttccccga gggcttcaag
      961 tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca ggactcctcc
     1021 ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt cccctccgac
     1081 ggccccgtaa tgcagaagaa gaccatgggc tgggaggcct cctccgagcg gatgtacccc
     1141 gaggacggcg ccctgaaggg cgagatcaag cagaggctga agctgaagga cggcggccac
     1201 tacgacgctg aggtcaagac cacctacaag gccaagaagc ccgtgcagct gcccggcgcc
     1261 tacaacgtca acatcaagtt ggacatcacc tcccacaacg aggactacac catcgtggaa
     1321 cagtacgaac gcgccgaggg ccgccactcc accggcggca tggacgagct gtacaagtcc
     1381 ggaggggatc ggtggagctc caccggtggc ggccgctcta gagagaatct ttattttcag
     1441 ggcgccgcca aattcaaaga aaccgctgct gctaaattcg aacgccagca catggacagc
     1501 ggaggtggag gtactagtct cgagatgagt gctgcagtga ctgcagggaa gctggcacgg
     1561 gcaccggccg accctgggaa agccggggtc cccggagttg cagctcccgg agctccggcg
     1621 gcggctccac cggcgaaaga gatcccggag gtcctagtgg acccacgcag ccggcggcgc
     1681 tatgtgcggg gccgcttttt gggcaagggc ggctttgcca agtgcttcga gatctcggac
     1741 gcggacacca aggaggtgtt cgcgggcagg attgtgccta agtctctgct gctcaagccg
     1801 caccagaggg agaagatgtc catggaaata tccattcacc gcagcctcgc ccaccagcac
     1861 gtcgtaggat tccacggctt tttcgaggac aacgacttcg tgttcgtggt gttggagctc
     1921 tgccgccgga ggtctctcct ggagctgcac aagaggagga aagccctgac tgagcctgag
     1981 gcccgatact acctacggca aattgtgctt ggctgccagt acctgcaccg aaaccgagtt
     2041 attcatcgag acctcaagct gggcaacctt ttcctgaatg aagatctgga ggtgaaaata
     2101 ggggattttg gactggcaac caaagtcgaa tatgacgggg agaggaagaa gaccctgtgt
     2161 gggactccta attacatagc tcccgaggtg ctgagcaaga aagggcacag tttcgaggtg
     2221 gatgtgtggt ccattgggtg tatcatgtat accttgttag tgggcaaacc accttttgag
     2281 acttcttgcc taaaagagac ctacctccgg atcaagaaga atgaatacag tattcccaag
     2341 cacatcaacc ccgtggccgc ctccctcatc cagaagatgc ttcagacaga tcccactgcc
     2401 cgcccaacca ttaacgagct gcttaatgac gagttcttta cttctggcta tatccctgcc
     2461 cgtctcccca tcacctgcct gaccattcca ccaaggtttt cgattgctcc cagcagcctg
     2521 gaccccagca accggaagcc cctcacagtc ctcaataaag gcttggagaa ccccctgcct
     2581 gagcgtcccc gggaaaaaga agaaccagtg gttcgagaga caggtgaggt ggtcgactgc
     2641 cacctcagtg acatgctgca gcagctgcac agtgtcaatg cctccaagcc ctcggagcgt
     2701 gggctggtca ggcaagagga ggctgaggat cctgcctgca tccccatctt ctgggtcagc
     2761 aagtgggtgg actattcgga caagtacggc cttgggtatc agctctgtga taacagcgtg
     2821 ggggtgctct tcaatgactc aacacgcctc atcctctaca atgatggtga cagcctgcag
     2881 tacatagagc gtgacggcac tgagtcctac ctcaccgtga gttcccatcc caactccttg
     2941 atgaagaaga tcaccctcct taaatatttc cgcaattaca tgagcgagca cttgctgaag
     3001 gcaggtgcca acatcacgcc gcgcgaaggt gatgagctcg cccggctgcc ctacctacgg
     3061 acctggttcc gcacccgcag cgccatcatc ctgcacctca gcaacggcag cgtgcagatc
     3121 aacttcttcc aggatcacac caagctcatc ttgtgcccac tgatggcagc cgtgacctac
     3181 atcgacgaga agcgggactt ccgcacatac cgcctgagtc tcctggagga gtacggctgc
     3241 tgcaaggagc tggccagccg gctccgctac gcccgcacta tggtggacaa gctgctgagc
     3301 tcacgctcgg ccagcaaccg tctcaaggcc tcctaaaagc ttcgaattct gcagtcgacg
     3361 gtaccgcggg cccgggatcc accggatcta gataactgat cataatcagc cataccacat
     3421 ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac ctgaaacata
     3481 aaatgaatgc aattgttgtt gttaacttgt ttattgcagc ttataatggt tacaaataaa
     3541 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     3601 tgtccaaact catcaatgta tcttaacgcg taaattgtaa gcgttaatat tttgttaaaa
     3661 ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa
     3721 atcccttata aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac
     3781 aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag
     3841 ggcgatggcc cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt
     3901 aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg
     3961 gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca
     4021 agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag
     4081 ggcgcgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     4141 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     4201 tattgaaaaa ggaagagtcc tgaggcggaa agaaccagct gtggaatgtg tgtcagttag
     4261 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     4321 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     4381 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     4441 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     4501 aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     4561 gcctaggctt ttgcaaagat cgatcaagag acaggatgag gatcgtttcg catgattgaa
     4621 caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac
     4681 tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg
     4741 cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaagacgag
     4801 gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt
     4861 gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg
     4921 tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg
     4981 catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga
     5041 gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag
     5101 gggctcgcgc cagccgaact gttcgccagg ctcaaggcga gcatgcccga cggcgaggat
     5161 ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt
     5221 tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg
     5281 gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt
     5341 tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc
     5401 ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac
     5461 gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg
     5521 acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccta
     5581 gggggaggct aactgaaaca cggaaggaga caataccgga aggaacccgc gctatgacgg
     5641 caataaaaag acagaataaa acgcacggtg ttgggtcgtt tgttcataaa cgcggggttc
     5701 ggtcccaggg ctggcactct gtcgataccc caccgagacc ccattggggc caatacgccc
     5761 gcgtttcttc cttttcccca ccccaccccc caagttcggg tgaaggccca gggctcgcag
     5821 ccaacgtcgg ggcggcaggc cctgccatag cctcaggtta ctcatatata ctttagattg
     5881 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     5941 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     6001 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     6061 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     6121 aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg tagccgtagt
     6181 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     6241 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     6301 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     6361 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     6421 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     6481 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     6541 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     6601 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     6661 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc atgcat
//
