LOCUS       sequence-467874-        6125 bp    DNA     circular SYN 15-APR-2025
DEFINITION  mammalian expression (CMV promoter) of  nuclear targeted BioID2-HA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6125)
  TITLE     The TDG Protein Environment Connects Active DNA Demethylation with 
            Chromatin and RNA Biology
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6125)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pCMV-NLS-BioID2-HA
FEATURES             Location/Qualifiers
     source          1..6125
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(46..65)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        237..616
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        617..820
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     771..791
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     865..884
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        865..883
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             909..929
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             966..1661
                     /codon_start=1
                     /product="promiscuous R40G mutant of a biotin protein 
                     ligase from Aquifex aeolicus (Kim et al., 2016)"
                     /label=BioID2
                     /translation="FKNLIWLKEVDSTQERLKEWNVSYGTALVADRQTKGRGGLGRKWL
                     SQEGGLYFSFLLNPKEFENLLQLPLVLGLSVSEALEEITEIPFSLKWPNDVYFQEKKVS
                     GVLCELSKDKLIVGIGINVNQREIPEEIKDRATTLYEITGKDWDRKEVLLKVLKRISEN
                     LKKFKEKSFKEFKGKIESKMLYLGEEVKLLGEGKITGKLVGLSEKGGALILTEEGIKEI
                     LSGEFSLRRS"
     CDS             1662..1688
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(1718..1735)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1724..1948
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1994..2422
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2081..2100)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2291..2312
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2431..2451)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2436..2765
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2616..2751
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2678..2697
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2832..3626
                     /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(2886..2905)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3496..3515
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3800..3921
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3837..3856)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3891..3910
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3970..3986)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3970..3986)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3983..4005)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3994..4010
                     /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(4018..4024,4025..4042,4043..4048))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4063..4084
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4201..4218)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4372..4960)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4452..4471)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5131..5922,5923..5991))
                     /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     5754..5773
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5992..6096)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga
       61 tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg
      121 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct
      181 gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca
      241 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata
      301 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
      361 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
      421 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
      481 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
      541 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
      601 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
      661 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
      721 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
      781 cggtaggcgt gtacggtggg aggtctatat aagcagagct ctctggctaa ctagagaacc
      841 cactgcttac tggcttatcg aaattaatac gactcactat agggagaccc aagctggcta
      901 gcatgggacc aaagaagaag cggaaggtcc accggtcgta cgtctccgga ttcgaattcg
      961 gatccttcaa gaacctgatc tggctgaagg aggtggacag cacccaggag agactgaagg
     1021 agtggaacgt gagctacggc accgccctgg tggccgacag acagaccaag ggcagaggcg
     1081 gcctgggcag aaagtggctg agccaggagg gcggcctgta cttcagcttc ctgctgaacc
     1141 ccaaggagtt cgagaacctg ctgcagctgc ccctggtgct gggcctgagc gtgagcgagg
     1201 ccctggagga gatcaccgag atccccttca gcctgaagtg gcccaacgac gtgtacttcc
     1261 aggagaagaa ggtgagcggc gtgctgtgcg agctgagcaa ggacaagctg atcgtgggca
     1321 tcggcatcaa cgtgaaccag agagagatcc ccgaggagat caaggacaga gccaccaccc
     1381 tgtacgagat caccggcaag gactgggaca gaaaggaggt gctgctgaag gtgctgaaga
     1441 gaatcagcga gaacctgaag aagttcaagg agaagagctt caaggagttc aagggcaaga
     1501 tcgagagcaa gatgctgtac ctgggcgagg aggtgaagct gctgggcgag ggcaagatca
     1561 ccggcaagct ggtgggcctg agcgagaagg gcggcgccct gatcctgacc gaggagggca
     1621 tcaaggagat cctgagcggc gagttcagcc tgagaagaag ctacccttac gatgtaccgg
     1681 attacgcata gaagcttaag tttaaaccgc tgatcagcct cgactgtgcc ttctagttgc
     1741 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     1801 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     1861 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     1921 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct
     1981 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     2041 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     2101 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     2161 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     2221 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     2281 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
     2341 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     2401 taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt
     2461 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     2521 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     2581 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     2641 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     2701 cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     2761 gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag acaggatgag
     2821 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     2881 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     2941 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     3001 tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
     3061 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     3121 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     3181 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     3241 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     3301 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc
     3361 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     3421 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     3481 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
     3541 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     3601 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc
     3661 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     3721 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     3781 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa
     3841 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     3901 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc
     3961 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     4021 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     4081 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     4141 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     4201 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     4261 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     4321 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     4381 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     4441 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     4501 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     4561 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     4621 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     4681 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     4741 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     4801 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     4861 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     4921 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     4981 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     5041 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     5101 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     5161 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     5221 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     5281 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     5341 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     5401 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     5461 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     5521 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     5581 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     5641 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     5701 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     5761 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     5821 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     5881 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     5941 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     6001 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     6061 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     6121 tgacg
//
