LOCUS       sequence-467886-        5850 bp    DNA     circular SYN 15-APR-2025
DEFINITION  mammalian expression (murine Pgk promoter) of nuclear targeted 
            BioID2-HA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5850)
  TITLE     The TDG Protein Environment Connects Active DNA Demethylation with 
            Chromatin and RNA Biology
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5850)
  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            "Unpublished"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: pPgk-NLS-BioID2-HA
FEATURES             Location/Qualifiers
     source          1..5850
                     /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"
     primer_bind     complement(204..227)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     554..573
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             634..654
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             691..1386
                     /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             1387..1413
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(1443..1460)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1449..1673
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1719..2147
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1806..1825)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2016..2037
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2156..2176)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2161..2490
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2341..2476
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2403..2422
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2557..3351
                     /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(2611..2630)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3221..3240
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3525..3646
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3562..3581)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3616..3635
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3695..3711)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3695..3711)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3708..3730)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3719..3735
                     /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(3743..3749,3750..3767,3768..3773))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3788..3809
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3926..3943)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4097..4685)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4177..4196)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4856..5647,5648..5716))
                     /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     5479..5498
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5717..5821)
                     /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 gacaattggg gtaggggagg
      181 cgcttttccc aaggcagtct ggagcatgcg ctttagcagc cccgctgggc acttggcgct
      241 acacaagtgg cctctggcct cgcacacatt ccacatccac cggtaggcgc caaccggctc
      301 cgttctttgg tggccccttc gcgccacctt ctactcctcc cctagtcagg aagttccccc
      361 ccgccccgca gctcgcgtcg tgcaggacgt gacaaatgga agtagcacgt ctcactagtc
      421 tcgtgcagat ggacagcacc gctgagcaat ggaagcgggt aggcctttgg ggcagcggcc
      481 aatagcagct ttgctccttc gctttctggg ctcaggggcg gggcgggcgc ccgaaggtcc
      541 tccggaggcc cggcattctg cacgcttcaa aagcgcacgt ctgccgcgct gttctcctct
      601 tcctcatctc cgggcctttc ggctagcatg ggaccaaaga agaagcggaa ggtccaccgg
      661 tcgtacgtct ccggattcga attcggatcc ttcaagaacc tgatctggct gaaggaggtg
      721 gacagcaccc aggagagact gaaggagtgg aacgtgagct acggcaccgc cctggtggcc
      781 gacagacaga ccaagggcag aggcggcctg ggcagaaagt ggctgagcca ggagggcggc
      841 ctgtacttca gcttcctgct gaaccccaag gagttcgaga acctgctgca gctgcccctg
      901 gtgctgggcc tgagcgtgag cgaggccctg gaggagatca ccgagatccc cttcagcctg
      961 aagtggccca acgacgtgta cttccaggag aagaaggtga gcggcgtgct gtgcgagctg
     1021 agcaaggaca agctgatcgt gggcatcggc atcaacgtga accagagaga gatccccgag
     1081 gagatcaagg acagagccac caccctgtac gagatcaccg gcaaggactg ggacagaaag
     1141 gaggtgctgc tgaaggtgct gaagagaatc agcgagaacc tgaagaagtt caaggagaag
     1201 agcttcaagg agttcaaggg caagatcgag agcaagatgc tgtacctggg cgaggaggtg
     1261 aagctgctgg gcgagggcaa gatcaccggc aagctggtgg gcctgagcga gaagggcggc
     1321 gccctgatcc tgaccgagga gggcatcaag gagatcctga gcggcgagtt cagcctgaga
     1381 agaagctacc cttacgatgt accggattac gcatagaagc ttaagtttaa accgctgatc
     1441 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     1501 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     1561 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     1621 ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga
     1681 ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
     1741 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     1801 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     1861 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     1921 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     1981 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     2041 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     2101 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat
     2161 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     2221 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     2281 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     2341 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     2401 tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga
     2461 ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc
     2521 ggatctgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac
     2581 gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca
     2641 atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt
     2701 gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg
     2761 tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga
     2821 agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct
     2881 cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg
     2941 gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg
     3001 gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc
     3061 gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat
     3121 ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac
     3181 tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt
     3241 gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct
     3301 cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc
     3361 tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca
     3421 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga
     3481 tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag
     3541 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     3601 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac
     3661 cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
     3721 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
     3781 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     3841 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     3901 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     3961 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     4021 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     4081 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     4141 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     4201 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     4261 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     4321 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     4381 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     4441 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     4501 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     4561 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     4621 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     4681 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     4741 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     4801 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     4861 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     4921 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     4981 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     5041 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     5101 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     5161 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     5221 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     5281 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     5341 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     5401 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     5461 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     5521 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     5581 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     5641 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     5701 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     5761 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     5821 gcacatttcc ccgaaaagtg ccacctgacg
//
