LOCUS       sequence-369089-        8351 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  RNAP2 Ser2ph-mintbody [42B3(R78K/A80T/M95V)-scFv] expression vector 
            for mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8351)
  AUTHORS   Uchino S, Ito Y, Sato Y, Handa T, Ohkawa Y, Tokunaga M, Kimura H
  TITLE     Live imaging of transcription sites using an elongating RNA 
            polymerase II-specific probe.
  JOURNAL   J Cell Biol. 2022 Feb 7;221(2). pii: 212888. doi: 
            10.1083/jcb.202104134. Epub 2021 Dec 2.
  PUBMED    34854870
REFERENCE   2  (bases 1 to 8351)
  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 Cell 
            Biol."; date: "2022-02-7"; volume: "221(2). pii: 212888. doi"; 
            pages: " 10.1083/jcb.202104134. Epub 2021 Dec 2"
COMMENT     Sequence Label: PB533-42B3mutAC2-sfGFP
FEATURES             Location/Qualifiers
     source          1..8351
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(1..105)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(131..586)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(268..289)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     480..499
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     713..735
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     727..744
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     728..744
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   874..936
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        1389..1600
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             1613..1881
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     regulatory      2671..2680
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2677..2679,2680..2682,2683..3396)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLL
                     EFVTAAGITHGMDELYK"
     primer_bind     complement(2722..2743)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     3330..3351
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    join(3406..3980,3981..3983,3984..3992)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3573..3590)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3800..3819
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4005..4799
                     /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
                     VLFVKTDLSGALNELQDEAARLSWLAATGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4059..4078)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4669..4688
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4859..4879)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    4921..5042
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4958..4977)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5012..5031
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   5805..5839
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(6330..6346)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6330..6346)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6343..6365)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6354..6370
                     /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(6378..6384,6385..6402,6403..6408))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6423..6444
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6561..6578)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6732..7320)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6812..6831)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7491..8282,8283..8351))
                     /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     8114..8133
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
       61 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
      121 agtgccacct aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
      181 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
      241 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
      301 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
      361 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
      421 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
      481 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
      541 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc
      601 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
      661 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca
      721 cgacgttgta aaacgacggc cagtgagcgc gcctcgttca ttcacgtttt tgaacccgtg
      781 gaggacgggc agactcgcgg tgcaaatgtg ttttacagcg tgatggagca gatgaagatg
      841 ctcgacacgc tgcagaacac gcagctagat taaccctaga aagataatca tattgtgacg
      901 tacgttaaag ataatcatgt gtaaaattga cgcatgtgtt ttatcggtct gtatatcgag
      961 gtttatttat taatttgaat agatattaag ttttattata tttacactta catactaata
     1021 ataaattcaa caaacaattt atttatgttt atttatttat taaaaaaaac aaaaactcaa
     1081 aatttcttct ataaagtaac aaaactttta tgagggacag ccccccccca aagcccccag
     1141 ggatgtaatt acgtccctcc cccgctaggg ggcagcagcg agccgcccgg ggctccgctc
     1201 cggtccggcg ctccccccgc atccccgagc cggcagcgtg cggggacagc ccgggcacgg
     1261 ggaaggtggc acgggatcgc tttcctctga acgcttctcg ctgctctttg agcctgcaga
     1321 cacctggggg gatacgggga aaaggcctcc acggccaagg atctgcgatc gctccggtgc
     1381 ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg
     1441 caattgaacg ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta
     1501 ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt
     1561 gaacgttctt tttcgcaacg ggtttgccgc cagaacacag ctgaagcttc gaggggctcg
     1621 catctctcct tcacgcgccc gccgccctac ctgaggccgc catccacgcc ggttgagtcg
     1681 cgttctgccg cctcccgcct gtggtgcctc ctgaactgcg tccgccgtct aggtaagttt
     1741 aaagctcagg tcgagaccgg gcctttgtcc ggcgctccct tggagcctac ctagactcag
     1801 ccggctctcc acgctttgcc tgaccctgct tgctcaactc tacgtctttg tttcgttttc
     1861 tgttctgcgc cgttacagat ccaagctgtg accggcgcct actctagagc tagcgaattc
     1921 gccatggccg aagtgcagct ggtggagtct gggggaggct tagtgaagcc tggagggtcc
     1981 ctgaaactct cctgtgcagc ctctggattc actttcagta gctttgccat gtcttgggtt
     2041 cgccagactc cggagaagag gctggagtgg gtcgcaacca ttagtagaag tggtagttac
     2101 acctactatt cagacagtgt gaagggtcga ttcaccatct ccagagacaa tgccaagaac
     2161 accctgtacc tgcaaatgag cagtctgagg tctgaggaca cggccgtcta ttactgtgca
     2221 agagacccct tcgatgtctg gggcgcaggg accacggtca ccgtctcctc aggtggaggc
     2281 ggttcaggcg gaggtggctc tggcggtggc ggatcgcagg ctgttgtgac tcaggaatct
     2341 gcactcacca catcacctgg tgaaacagtc acactcactt gtcgctcaag tactggggct
     2401 gttacaacta gtaactatgc caactgggtc caagaaaaac cagatcattt attcactggt
     2461 ctaataggtg gtaccaacaa ccgagctcca ggtgttcctg ccagattctc aggctccctg
     2521 attggagaca aggctgccct caccatcaca ggggcacaga ctgaggatga ggcaatatat
     2581 ttctgtgctc tatggtacag caacctttgg gtgttcggtg gaggaaccaa actgactgtc
     2641 ctaggccagc ccaaggcgga tccaccggtc gccaccatgg tgagcaaggg cgaggagctg
     2701 ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc
     2761 agcgtgcgcg gcgagggcga gggcgatgcc accaacggca agctgaccct gaagttcatc
     2821 tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctacggc
     2881 gtgcagtgct tcagccgcta ccccgaccac atgaagcgcc acgacttctt caagtccgcc
     2941 atgcccgaag gctacgtcca ggagcgcacc atcagcttca aggacgacgg cacctacaag
     3001 acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc
     3061 atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa cttcaacagc
     3121 cacaacgtct atatcaccgc cgacaagcag aagaacggca tcaaggccaa cttcaagatc
     3181 cgccacaacg tggaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc
     3241 atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca gtccgtgctg
     3301 agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc
     3361 gggatcactc acggcatgga cgagctgtac aagtaagcgg ccgcgcccct ctccctcccc
     3421 cccccctaac gttactggcc gaagccgctt ggaataaggc cggtgtgcgt ttgtctatat
     3481 gttattttcc accatattgc cgtcttttgg caatgtgagg gcccggaaac ctggccctgt
     3541 cttcttgacg agcattccta ggggtctttc ccctctcgcc aaaggaatgc aaggtctgtt
     3601 gaatgtcgtg aaggaagcag ttcctctgga agcttcttga agacaaacaa cgtctgtagc
     3661 gaccctttgc aggcagcgga accccccacc tggcgacagg tgcctctgcg gccaaaagcc
     3721 acgtgtataa gatacacctg caaaggcggc acaaccccag tgccacgttg tgagttggat
     3781 agttgtggaa agagtcaaat ggctctcctc aagcgtattc aacaaggggc tgaaggatgc
     3841 ccagaaggta ccccattgta tgggatctga tctggggcct cggtgcacat gctttacatg
     3901 tgtttagtcg aggttaaaaa aacgtctagg ccccccgaac cacggggacg tggttttcct
     3961 ttgaaaaaca cgatgataat atggccacaa ccatggcgtc cggaatgatt gaacaagatg
     4021 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
     4081 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
     4141 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc
     4201 ggctatcgtg gctggccgcg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
     4261 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     4321 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     4381 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     4441 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     4501 cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg
     4561 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     4621 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     4681 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     4741 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag
     4801 tcgacaatca acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg
     4861 ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gcgttaacta
     4921 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     4981 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     5041 tatcatgtct ggaattgact caaatgatgt caattagtct atcagaagct catctggtct
     5101 cccttccggg ggacaagaca tccctgttta atatttaaac agcagtgttc ccaaactggg
     5161 ttcttatatc ccttgctctg gtcaaccagg ttgcagggtt tcctgtcctc acaggaacga
     5221 agtccctaaa gaaacagtgg cagccaggtt tagccccgga attgactgga ttcctttttt
     5281 agggcccatt ggtatggctt tttccccgta tccccccagg tgtctgcagg ctcaaagagc
     5341 agcgagaagc gttcagagga aagcgatccc gtgccacctt ccccgtgccc gggctgtccc
     5401 cgcacgctgc cggctcgggg atgcgggggg agcgccggac cggagcggag ccccgggcgg
     5461 ctcgctgctg ccccctagcg ggggagggac gtaattacat ccctgggggc tttggggggg
     5521 ggctgtccct gatatctata acaagaaaat atatatataa taagttatca cgtaagtaga
     5581 acatgaaata acaatataat tatcgtatga gttaaatctt aaaagtcacg taaaagataa
     5641 tcatgcgtca ttttgactca cgcggtcgtt atagttcaaa atcagtgaca cttaccgcat
     5701 tgacaagcac gcctcacggg agctccaagc ggcgactgag atgtcctaaa tgcacagcga
     5761 cggattcgcg ctatttagaa agagagagca atatttcaag aatgcatgcg tcaattttac
     5821 gcagactatc tttctagggt taatctagct gcatcaggat catatcgtcg ggtctttttt
     5881 ccggctcagt catcgcccaa gctggcgcta tctgggcatc ggggaggaag aagcccgtgc
     5941 cttttcccgc gaggttgaag cggcatggaa agagtttgcc gaggatgact gctgctgcat
     6001 tgacgttgag cgaaaacgca cgtttaccat gatgattcgg gaaggtgtgg ccatgcacgc
     6061 ctttaacggt gaactgttcg ttcaggccac ctgggatacc agttcgtcgc ggcttttccg
     6121 gacacagttc cggatggtca gcccgaagcg catcagcaac ccgaacaata ccggcgacag
     6181 ccggaactgc cgtgccggtg tgcagattaa tgacagcggt gcggcgctgg gatattacgt
     6241 cagcgaggac gggtatcctg gctggatgcc gcagaaatgg acatggatac cccgtgagtt
     6301 acccggcggg cgcgcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     6361 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     6421 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     6481 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     6541 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     6601 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     6661 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     6721 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     6781 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     6841 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     6901 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     6961 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     7021 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     7081 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     7141 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     7201 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     7261 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     7321 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     7381 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     7441 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     7501 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     7561 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     7621 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     7681 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     7741 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     7801 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     7861 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     7921 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     7981 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     8041 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     8101 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     8161 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     8221 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     8281 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     8341 tgaatactca t
//
