LOCUS       sequence-467871-        6407 bp    DNA     circular SYN 15-APR-2025
DEFINITION  mammalian expression (CMV promoter) of histone H4 fused to 
            BioID2-HA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6407)
  TITLE     The TDG Protein Environment Connects Active DNA Demethylation with 
            Chromatin and RNA Biology
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6407)
  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            "Unpublished"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-H4-BioID2-HA
FEATURES             Location/Qualifiers
     source          1..6407
                     /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             1248..1943
                     /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             1944..1970
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(2000..2017)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2006..2230
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2276..2704
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2363..2382)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2573..2594
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2713..2733)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2718..3047
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2898..3033
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2960..2979
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3114..3908
                     /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(3168..3187)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3778..3797
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4082..4203
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4119..4138)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4173..4192
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4252..4268)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4252..4268)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4265..4287)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4276..4292
                     /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(4300..4306,4307..4324,4325..4330))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4345..4366
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4483..4500)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4654..5242)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4734..4753)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5413..6204,6205..6273))
                     /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     6036..6055
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6274..6378)
                     /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 gcatgtccgg cagaggaaag ggcggaaaag gcttaggcaa agggggcgct aagcgccacc
      961 gcaaggtctt gagagacaac attcagggca tcaccaagcc tgccattcgg cgtctagctc
     1021 ggcgtggcgg cgttaagcgg atctctggcc tcatttacga ggagacccgc ggtgtgctga
     1081 aggtgttcct ggagaatgtg attcgggacg cagtcaccta caccgagcac gccaagcgca
     1141 agaccgtcac agccatggat gtggtgtacg cgctcaagcg ccaggggcgc accctgtacg
     1201 gcttcggagg ctaccggtcg tacgtctccg gattcgaatt cggatccttc aagaacctga
     1261 tctggctgaa ggaggtggac agcacccagg agagactgaa ggagtggaac gtgagctacg
     1321 gcaccgccct ggtggccgac agacagacca agggcagagg cggcctgggc agaaagtggc
     1381 tgagccagga gggcggcctg tacttcagct tcctgctgaa ccccaaggag ttcgagaacc
     1441 tgctgcagct gcccctggtg ctgggcctga gcgtgagcga ggccctggag gagatcaccg
     1501 agatcccctt cagcctgaag tggcccaacg acgtgtactt ccaggagaag aaggtgagcg
     1561 gcgtgctgtg cgagctgagc aaggacaagc tgatcgtggg catcggcatc aacgtgaacc
     1621 agagagagat ccccgaggag atcaaggaca gagccaccac cctgtacgag atcaccggca
     1681 aggactggga cagaaaggag gtgctgctga aggtgctgaa gagaatcagc gagaacctga
     1741 agaagttcaa ggagaagagc ttcaaggagt tcaagggcaa gatcgagagc aagatgctgt
     1801 acctgggcga ggaggtgaag ctgctgggcg agggcaagat caccggcaag ctggtgggcc
     1861 tgagcgagaa gggcggcgcc ctgatcctga ccgaggaggg catcaaggag atcctgagcg
     1921 gcgagttcag cctgagaaga agctaccctt acgatgtacc ggattacgca tagaagctta
     1981 agtttaaacc gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc
     2041 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa
     2101 aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg
     2161 gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg
     2221 ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg
     2281 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     2341 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     2401 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     2461 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     2521 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     2581 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     2641 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     2701 aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag
     2761 gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag
     2821 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc
     2881 cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc
     2941 atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc tctgagctat
     3001 tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcccgggag
     3061 cttgtatatc cattttcgga tctgatcaag agacaggatg aggatcgttt cgcatgattg
     3121 aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg
     3181 actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg
     3241 ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg
     3301 aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg
     3361 ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc
     3421 tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc
     3481 tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc
     3541 gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc
     3601 aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg
     3661 atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct
     3721 tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt
     3781 tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc
     3841 tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt
     3901 tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc
     3961 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     4021 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     4081 caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     4141 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     4201 ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct
     4261 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     4321 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     4381 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     4441 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     4501 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     4561 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     4621 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     4681 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     4741 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     4801 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     4861 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     4921 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     4981 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     5041 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     5101 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     5161 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     5221 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     5281 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     5341 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     5401 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     5461 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     5521 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     5581 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     5641 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     5701 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     5761 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     5821 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     5881 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     5941 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     6001 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     6061 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     6121 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     6181 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     6241 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     6301 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     6361 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacg
//
