LOCUS       sequence-94262-e        6912 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express Flag-tagged RCC1 in mammalian cells (CMV promoter). Also 
            suitable for in vitro transcription (T7 promoter) for injecting mRNA
            into oocytes..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6912)
  AUTHORS   Inoue A, Zhang Y
  TITLE     Nucleosome assembly is required for nuclear pore complex assembly in
            mouse zygotes.
  JOURNAL   Nat Struct Mol Biol. 2014 Jul;21(7):609-16. doi: 10.1038/nsmb.2839. 
            Epub 2014 Jun 8.
  PUBMED    24908396
REFERENCE   2  (bases 1 to 6912)
  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: "10.1038/nsmb.2839";
            journalName: "Nat Struct Mol Biol"; date: "2014-07"; volume: "21"; 
            issue: "7"; pages: "609-16"
COMMENT     Sequence Label: pcDNA-Flag-RCC1-polyA
FEATURES             Location/Qualifiers
     source          1..6912
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             911..934
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2415..2444
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2460..2477
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2500..2517)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2506..2730
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2776..3204
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2863..2882)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3073..3094
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3213..3233)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3218..3548
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3399..3534
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3461..3480
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3615..4409
                     /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(3669..3688)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4279..4298
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4585..4706
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4622..4641)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4676..4695
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4755..4771)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4755..4771)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4768..4790)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4779..4795
                     /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(4803..4809,4810..4827,4828..4833))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4848..4869
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4986..5003)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5157..5745)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5237..5256)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5916..6707,6708..6776))
                     /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     6539..6558
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6777..6881)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagt
      901 taagcttatg gactacaaag acgatgacga caagggatcc ccacccaagc gcatagctaa
      961 gagaaggtca cccccagaag atgccatccc caaaagcaag aaggtgaaag tctcacacag
     1021 gtcccacaac acagaaccag gcttggtgct gacactgggc cagggcgacg tgggccagct
     1081 ggggctgggt gaaagtgtgc tggagaggaa gaagccggcc ttggtgcccc ttctgcagga
     1141 tgttgtgcag gctgaggccg ggggcatgca tactgtgtgt ctgagccaaa gtggccaggt
     1201 ctactccttt ggctgcaatg acgagggtgc cctgggaagg gacacttcag tcgaaggctc
     1261 agaaatggtc cctgggaaag tggaactgca agagaaagtg gtgcaagtgt cagcggggga
     1321 cagtcacaca gcagccctca ccgaagacgg ccgtgtcttc ctttggggct ctttccggga
     1381 caataatggt gtgatcgggt tgttggagcc catgaagaag agcatggtac ctgttcaagt
     1441 gcagctggat gcgcccgtgg taaaggtggc ctcggggaac gaccacttgg tgatgctcac
     1501 gaacgatggc gacctctaca ccttgggttg tggggagcaa ggtcagctgg gccgtgtacc
     1561 tgaattattt gccaaccgag gtggccgtca gggccttggg cgactcctgg tccccagatg
     1621 tgtgctgctg aaatccaggg ggacccgggg ccgtgtgcgg ttccaggatg ccttctgtgg
     1681 cgcctatttc acttttgcca tctcccgaga gggccacgtg tatggctttg gcctctccaa
     1741 ctatcaccag ctcgggactc caggcactgg atcttgcttc attcctcaaa acttgacatc
     1801 cttcaagaat tccaccaagt cctgggtagg cttctccggt ggccagcatc acacagtctg
     1861 catggattcg gaagggaagg cgtacagcct gggccgggct gagtacgggc gcctgggcct
     1921 tggggagggt gctgaggaga agagcatacc caccctcatt tcccggctgc ccgtggtctc
     1981 ctcagtggcc tgtggggctt ctgtgggtta tgctgtgtcc aaggatggtc gtgtttttgc
     2041 ctggggcatg ggcaccaact accagctggg caccgggcag gatgaagatg cctggagtcc
     2101 tgtggaaatg actggcaaac agctggagaa ccgtgtggtg ttaactgtgt ccagtggggg
     2161 ccagcacacg gtcttactag ttaaggacca ggcacagagc tgagcggccg ctcgactatg
     2221 ggcaccaaag aacctgtaaa cgttatcttt ttaaattgaa tgtgcacaaa taaaagtttg
     2281 gaaaagaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa
     2341 aaaaaaaaaa aaaaaaaaaa aaaaaaaaat acgcatcata cgctgtggcg tacctcgagt
     2401 ctagagggcc cttcgaacaa aaactcatct cagaagagga tctgaatatg cataccggtc
     2461 atcatcacca tcaccattga gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt
     2521 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     2581 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     2641 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
     2701 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct
     2761 ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     2821 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     2881 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg ggcatccctt
     2941 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     3001 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     3061 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
     3121 attcttttga tttataaggg attttgggga tttcggccta ttggttaaaa aatgagctga
     3181 tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa
     3241 gtccccaggc tccccaggca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     3301 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     3361 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     3421 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     3481 ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     3541 tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcaa gagacaggat
     3601 gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg
     3661 tggagaggct attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg
     3721 tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg
     3781 ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc
     3841 cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg
     3901 aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca
     3961 tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc
     4021 aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg
     4081 atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg
     4141 cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata
     4201 tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg
     4261 accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat
     4321 gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct
     4381 tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgcga aatgaccgac
     4441 caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg
     4501 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     4561 atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa
     4621 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     4681 ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc
     4741 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     4801 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     4861 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     4921 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     4981 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     5041 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     5101 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     5161 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     5221 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     5281 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     5341 gcgctttctc aatgctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     5401 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     5461 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     5521 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     5581 tacggctaca ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     5641 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     5701 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     5761 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     5821 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     5881 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     5941 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     6001 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     6061 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     6121 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     6181 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     6241 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     6301 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     6361 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     6421 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     6481 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     6541 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     6601 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     6661 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     6721 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     6781 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     6841 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     6901 ccacctgacg tc
//
