LOCUS       sequence-65993-e        7838 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 7838)
  AUTHORS   Kiyomitsu T, Cheeseman IM
  TITLE     Cortical dynein and asymmetric membrane elongation coordinately 
            position the spindle in anaphase.
  JOURNAL   Cell. 2013 Jul 18;154(2):391-402. doi: 10.1016/j.cell.2013.06.010.
  PUBMED    23870127
REFERENCE   2  (bases 1 to 7838)
  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: 
            "10.1016/j.cell.2013.06"; journalName: "Cell"; date: "2013-07-18- 
            18"; volume: "154"; issue: "2"; pages: "391-402"
COMMENT     Sequence Label: pTK162_mCherry-4.1G-hardened
FEATURES             Location/Qualifiers
     source          1..7838
                     /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"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             613..1320
                     /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(758..776)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1026..1045
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1366..1386
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1399..1443
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     polyA_signal    4626..4747
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4663..4682)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4717..4736
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4754..5209)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4891..4912)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5103..5122
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5236..5340
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5342..5699
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5342..5533
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5365..5385)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5550..5685
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5734..6528
                     /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             5734..6528
                     /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(5788..5807)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6398..6417
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6716..6735)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6760..6807
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      7136..7724
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7625..7644
                     /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 gctagcgcta
      601 ccggtcgcca ccatggtgag caagggcgag gaggataaca tggccatcat caaggagttc
      661 atgcgcttca aggtgcacat ggagggctcc gtgaacggcc acgagttcga gatcgagggc
      721 gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggt
      781 ggccccctgc ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg ctccaaggcc
      841 tacgtgaagc accccgccga catccccgac tacttgaagc tgtccttccc cgagggcttc
      901 aagtgggagc gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc
      961 tccctgcagg acggcgagtt catctacaag gtgaagctgc gcggcaccaa cttcccctcc
     1021 gacggccccg taatgcagaa gaagaccatg ggctgggagg cctcctccga gcggatgtac
     1081 cccgaggacg gcgccctgaa gggcgagatc aagcagaggc tgaagctgaa ggacggcggc
     1141 cactacgacg ctgaggtcaa gaccacctac aaggccaaga agcccgtgca gctgcccggc
     1201 gcctacaacg tcaacatcaa gttggacatc acctcccaca acgaggacta caccatcgtg
     1261 gaacagtacg aacgcgccga gggccgccac tccaccggcg gcatggacga gctgtacaag
     1321 tccggagggg atcggtggag ctccaccggt ggcggccgct ctagagagaa tctttatttt
     1381 cagggcgccg ccaaattcaa agaaaccgct gctgctaaat tcgaacgcca gcacatggac
     1441 agcggaggtg gaggtactag tctcgagatg actactgaag taggctctgt gtctgaagtg
     1501 aagaaggact ctagccagtt aggaacagat gcaaccaagg aaaaacctaa agaagtagca
     1561 gaaaatcagc agaatcagtc ttccgatcca gaggaggaaa aaggttccca gccacctcct
     1621 gcagctgaaa gccaaagtag tctacgccgc cagaagagag agaaggaaac atcggagagc
     1681 aggggtattt ctcggttcat accgccatgg cttaagaagc aaaagtcata taccttagta
     1741 gtggccaaag atggaggaga taaaaaagag cctacccaag ctgttgttga agaacaggtc
     1801 ttagataaag aggaacccct tccagaagaa cagagacagg ctaagggtga tgctgaagaa
     1861 atggctcaga agaaacaaga gattaaagtt gaagtcaagg aagaaaaacc ctcagtgagc
     1921 aaggaagaaa aaccctcagt gagcaaagtg gagatgcagc ctactgaatt agtaagtaag
     1981 gagagagaag agaaggtaaa agaaacacag gaagacaaat tagaaggagg agcagcaaaa
     2041 agggagacca aggaagtgca gaccaatgag ctgaaagcag agaaggcatc tcaaaaagtc
     2101 accaagaaga ccaaaactgt ccagtgtaaa gtgaccctct tagatggcac cgaatacagc
     2161 tgtgacctgg agaaacatgc caagggacaa gtgttatttg acaaagtgtg tgaacacctc
     2221 aatctcttgg agaaagacta ctttggactt ttgtttcagg aaagccctga gcagaaaaac
     2281 tggttagatc ctgctaaaga aataaagaga caactgagaa accttccatg gctattcact
     2341 tttaatgtga agttttatcc tcctgatcct tctcaattga ctgaagatat caccagatac
     2401 ttcttgtgcc ttcagctccg gcaggacatt gcctctggcc gcctgccctg ctcttttgtg
     2461 actcatgctc tcctgggatc ctacaccctg caggctgaac ttggtgacta tgacccagaa
     2521 gaacatggca gcatcgacct cagtgaattc cagtttgccc ctactcagac taaggagctg
     2581 gaagagaagg tggcagagct gcacaaaacc cacaggggct tatcgccagc acaagctgat
     2641 tcccagttct tagaaaatgc aaagaggctt tccatgtatg gtgttgacct acatcatgcc
     2701 aaggactcag aaggtgtgga catcaagctg ggcgtgtgtg ctaatggact tctcatttac
     2761 aaagacagac tgcgaatcaa tcgttttgct tggccgaaaa tcttaaaaat ttcctataaa
     2821 cgcagtaact tctacattaa agtcagaccg gcagagctgg aacagtttga gagtaccatt
     2881 ggattcaaac tgccaaacca ccgggcagcg aaaagactat ggaaagtgtg cgtggagcat
     2941 catactttct acaggcttgt ttctccagag cagccaccaa aagccaagtt cctgaccttg
     3001 gggtccaaat ttcgctatag tggccgcacc caagcacaga cccgccaggc cagcaccctc
     3061 atagataggc cagctcctca tttcgaacgg acaagcagta aacgggtctc caggagtcta
     3121 gatggagctc cgattggtgt catggaccaa agtcttatga aggattttcc tggcgctgct
     3181 ggggagattt cagcctatgg acctggactt gtcagcattg ccgtggtaca agatggggac
     3241 ggcaggaggg aagtgagaag cccaactaaa gccccacatt tgcagctcat tgaaggaaag
     3301 aaaaattcct tgagagtaga aggggataat atttatgtca gacatagcaa tttaatgttg
     3361 gaggaactgg ataaggccca ggaggacata ctgaaacatc aggctagcat tagtgaactc
     3421 aagcgcaatt ttatggaatc cacacctgag ccgcgcccta atgaatggga aaaacgacgt
     3481 atcacacctc tgtccctaca gacacaaggg agttcacatg agactctgaa tatagtggag
     3541 gagaagaagc gggcagaggt tgggaaagac gaaagagtaa tcacagaaga aatgaatggt
     3601 aaagagatat cacctgggag tggtcctggg gagattcgta aggtggagcc tgtgacacaa
     3661 aaagactcca cctccctgtc ttctgagagc agcagcagca gcagtgagag tgaggaggaa
     3721 gacgtgggag agtaccgtcc ccaccaccga gtgaccgagg gcaccatcag ggaggaacag
     3781 gagtatgaag aagaggtgga ggaagaaccc cgcccggcag ccaaggtagt agagagggag
     3841 gaagcagtgc ccgaagccag cccagtcaca caagcaggtg ccagtgtaat cacagtagaa
     3901 acagtgatcc aggaaaatgt aggtgcccaa aagatacccg gagagaagag tgtacacgaa
     3961 ggcgctctta agcaagacat gggagaagaa gcagaggaag agccacagaa agttaacgga
     4021 gaggtgtccc atgttgacat tgatgttttg ccacaaatta tttgttgttc agagccacca
     4081 gtggtaaaaa cagagatggt aacaatttct gatgcctcac aaaggacaga aatctccacc
     4141 aaggaagtcc ccattgtcca aactgagacc aaaaccatca catatgagtc tccacagatt
     4201 gatggcgggg ctggtggtga ttcgggcacg ttactgaccg cacaaaccat cacatctgag
     4261 tccgtgtcaa caacgacaac cacacacatc accaagactg taaaaggtgg aatttctgaa
     4321 acaagaattg agaaacgcat tgtgatcaca ggagatggag atattgatca tgaccaggca
     4381 ctggctcagg cgatcaggga agccagagag cagcaccctg acatgtcggt cacaagagtg
     4441 gtggtacaca aagaaacaga gttggctgag gaaggggaag attaaccgcg ggcccgggat
     4501 ccaccggatc tagataactg atcataatca gccataccac atttgtagag gttttacttg
     4561 ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg
     4621 ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     4681 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     4741 tatcttaacg cgtaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt
     4801 taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa
     4861 gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag
     4921 aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt
     4981 gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac
     5041 cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag
     5101 gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg
     5161 cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc aggtggcact
     5221 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     5281 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     5341 cctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
     5401 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
     5461 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     5521 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     5581 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg
     5641 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaag
     5701 atcgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca
     5761 ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc
     5821 ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc
     5881 aagaccgacc tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg
     5941 ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg
     6001 gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct
     6061 gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct
     6121 acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa
     6181 gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa
     6241 ctgttcgcca ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt gacccatggc
     6301 gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt
     6361 ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct
     6421 gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc
     6481 gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg
     6541 ggttcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg
     6601 ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc
     6661 tccagcgcgg ggatctcatg ctggagttct tcgcccaccc tagggggagg ctaactgaaa
     6721 cacggaagga gacaataccg gaaggaaccc gcgctatgac ggcaataaaa agacagaata
     6781 aaacgcacgg tgttgggtcg tttgttcata aacgcggggt tcggtcccag ggctggcact
     6841 ctgtcgatac cccaccgaga ccccattggg gccaatacgc ccgcgtttct tccttttccc
     6901 caccccaccc cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag
     6961 gccctgccat agcctcaggt tactcatata tactttagat tgatttaaaa cttcattttt
     7021 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     7081 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     7141 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     7201 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     7261 gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga
     7321 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     7381 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     7441 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     7501 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     7561 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     7621 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     7681 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     7741 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat
     7801 cccctgattc tgtggataac cgtattaccg ccatgcat
//
