LOCUS       sequence-491709-        7293 bp    DNA     circular SYN 04-NOV-2025
DEFINITION  Expresses HaloTag-TAF1A.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7293)
  AUTHORS   Ide S, Imai R, Ochi H, Maeshima K
  TITLE     Transcriptional suppression of ribosomal DNA with phase separation.
  JOURNAL   Sci Adv. 2020 Oct 14;6(42). pii: 6/42/eabb5953. doi: 
            10.1126/sciadv.abb5953. Print 2020 Oct.
  PUBMED    33055158
REFERENCE   2  (bases 1 to 7293)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 4, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Sci Adv."; 
            date: "2020-10-14"; volume: "6(42). pii: 6/42/eabb5953. doi"; pages:
            " 10.1126/sciadv.abb5953. Print 2020 Oct"
COMMENT     Sequence Label: pEF1-FRT-HaloTag-TAF1A
FEATURES             Location/Qualifiers
     source          1..7293
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     85..102
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(140..181)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             complement(220..1104)
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="EIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSYV
                     WRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIHD
                     WGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIIDQ
                     NVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVEE
                     YMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLIG
                     SEIARWLSTLEISG"
     regulatory      1871..1880
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      1875..1884
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1914..1933)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1915..1933)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1940..1957)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1940..1956)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1949..1971)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2097..2552
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2184..2203)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2394..2415
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2630..2734
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2735..2803,2804..3595)
                     /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     complement(2953..2972)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3766..4354
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4255..4274
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4508..4525
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4642..4663
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4678..4683,4684..4701,4702..4708)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4716..4732
                     /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)."
     primer_bind     4721..4743
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4740..4756
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4740..4756
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        4774..4792
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     4774..4791
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     polyA_signal    5757..5838
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     5782..5801
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(5937..6536)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(6020..6040)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     6517..6536
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(6558..7068)
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     complement(6621..6639)
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     polyA_signal    join(7165..7293,1..96)
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 tttattagga aaggacagtg ggagtggcac cttccagggt caaggaaggc acgggggagg
       61 ggcaaacaac agatggctgg caactagaag gcacagtcga ggctgatcag cgggtttaaa
      121 ctcattacta accggtacgc gtagaatcga gaccgaggag agggttaggg ataggcttac
      181 cttcgaaccg cgggccctct agactcgagc ggccgcttaa ccggaaatct ccagagtaga
      241 cagccagcgc gcgatctcgc tgccgatcag gtccgggttg tcttcttgca gcagattcag
      301 acccgggccg atgtccacag ccttgcagtt aggcaggctt ttggccaggc gagcggcttc
      361 ggccggtggg atcagaacgc ctggggtgcc ccagaacagc agcttcggga caggggactg
      421 gtgcagccag tccatgtatt cttcgaccag cgcgacgatg ttcgctggct caccggcgat
      481 tggcagctcg tttgggaagc gccacagtgg ctcgcggtca acaggattca ggaacggctc
      541 gcggtaatgg tccatctcga cttcagtcag cgggcggacg acacccatcg gcagcgtacc
      601 ctcgataaaa acgttctgat cgatgatcag cttgcggccg acgtcggtgg tgcggaaggc
      661 ctggaaggtc tcgcgggcaa attctggcca ttcgtcccag gtcgggatag ggcggatgaa
      721 ctccataaat gcaatacctt tgacgcgctc tggattgcgc ttggcccagt ggaaacccag
      781 agcggagccc cagtcgtgaa tgaccaggac gacctcttcc agacccaggg cttcgatgaa
      841 ggcatccatg aagcggacgt ggtcgtcgaa gaaataaccc aggtctggtt tgtcggattt
      901 gcccataccg atcaggtctg gagcaatgca gcgatgggtc ggtgcaacat gcgggatgat
      961 gttgcgccac acgtaggagg aggtcgggtt accgtgcagg aacagcacag gggtgccatc
     1021 gcgcggacca acatcgacgt agtgcatgcg ctcgcccagg acttccacat aatgggggtc
     1081 gaatggaaag ccagtaccga tttcggatcc cattgaaccg ccgccaccag atccacctcc
     1141 accagatcca cctccaccga gtcttggatt tacaatactg tattttttca cagatctctt
     1201 catccgctta attttcttcc ctaagatttg gtgatcttgc tttaaaatat accggaaata
     1261 tctacaacct aaaaagttaa gcaaaataaa aagttttaat actcatcaca tttttcaagt
     1321 atatttctac attttaaaaa ttctgcttta agtttagcac actaatgatt atgaaaatag
     1381 atttatttgt attatcaata tgtcctaagc atattctcag ctgtgcccaa tgaaatacat
     1441 ttgttcctat atacaaaatt tatttgtatt accaatatgg cataagtata ttctcagctg
     1501 tgcccaacga aacatatttc ttcctatata taaagatgtt tctaatccac cataattaaa
     1561 gcatagactt tagcttgttt tctactcttc ctcctacacc ctagtgatgg tgacaggcat
     1621 attatataca gtcacatatt aaatgagcct tctccaaaag cacattcgta tagcttcctt
     1681 ctcaatgtta gcttgtaaat cagccaaagt tacgttcata acagtaagaa agttctctgt
     1741 agaaaatctt aggatgtgaa ggattagaat aaacaaatca taggttagaa acggtcaaac
     1801 attatctatt cattctagcc atgtagagag cagcaaggtg gagaccatgt agctctcacg
     1861 aattcccgcg gccgccatgg cggccgggag catgcgacgt cgggcccaat tcgccctata
     1921 gtgagtcgta ttacaattca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg
     1981 gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg
     2041 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggacgc
     2101 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     2161 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     2221 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     2281 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     2341 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     2401 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     2461 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     2521 gaattttaac aaaatattaa cgcttacaat ttcctgatgc ggtattttct ccttacgcat
     2581 ctgtgcggta tttcacaccg catcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     2641 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     2701 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     2761 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     2821 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     2881 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     2941 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact
     3001 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     3061 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     3121 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     3181 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     3241 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg
     3301 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     3361 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     3421 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc
     3481 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     3541 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     3601 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     3661 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     3721 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     3781 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     3841 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     3901 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     3961 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     4021 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     4081 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     4141 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     4201 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     4261 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     4321 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     4381 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     4441 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac
     4501 cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct
     4561 ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc
     4621 gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt
     4681 acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac
     4741 aggaaacagc tatgaccatg attacgccaa gctatttagg tgacactata gaatactcaa
     4801 gctatgcatc caacgcgttg ggagctctcc catatggtcg acgcccactc tactgctatg
     4861 acccaaatta gtcctctaga gtcaaaggtc tctctctagc aggaggttat tttgccccaa
     4921 actgtcatgt ctttatactg aaagagtctt tttgtttttt tttttttttt tgagacagag
     4981 tctagttctg tcgcccaggc aggagtacgg tggtgcaatc tcggctcact gcaacctcca
     5041 cctcccgggt tcaagcaatt ctcctgcttc agcctcccga gtatctggga ttacaaatgc
     5101 ccgccactgc gcccagctac tttttgtatt tttagtagag acagggtttc actgtgttgg
     5161 ccaggctggt cttgaactcc tgacctcatg atccacccac ctcggcctcc caagtgctgg
     5221 gattagaagt gtgagccact gcgcccggcc gagaaagagt cttttttcta aatccttctc
     5281 tcttttcaaa attcctcaat agaactcatc accaatatat ttcttcatgg tggtatacct
     5341 aaaatggatt aattagaata taaaagtaga atataaatac atttcacagc actacaagtt
     5401 acagtttatt acacaatagc aagaggcaaa gtatttgcaa acaataactg ctgtagagct
     5461 atattctgta attagtttgt catttgaaga attattcctt aacaaatgta acatttaaca
     5521 ttacaaaaaa gaaaacagac atagttacaa agagcaaagg cagtaatatt gtttctctag
     5581 ctataaataa tacaaaaata taaaaatata taaaaataag ttttttgtag taatataagc
     5641 ttcttaatac actacataaa tacattcaat aactatctac caagctactt actgtgtagc
     5701 tacaactgtg aaataactaa aattcagtat cagtttggac aaaccacaac tagaatgcag
     5761 tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata
     5821 agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg
     5881 gaggtgtggg aggtttttta aagcaagtaa aacctctaca aatgtggtat ggctgatcag
     5941 gcaccgggct tgcgggtcat gcaccaggtg cgcggtcctt cgggcacctc gacgtcggcg
     6001 gtgacggtga agccgagccg ctcgtagaag gggaggttgc ggggcgcgga ggtctccagg
     6061 aaggcgggca ccccggcgcg ctcggccgcc tccactccgg ggagcacgac ggcgctgccc
     6121 agacccttgc cctggtggtc gggcgagacg ccgacggtgg ccaggaacca cgcgggctcc
     6181 ttgggccggt gcggcgccag gaggccttcc atctgttgct gcgcggccag ccgggaaccg
     6241 ctcaactcgg ccatgcgcgg gccgatctcg gcgaacaccg cccccgcttc gacgctctcc
     6301 ggcgtggtcc agaccgccac cgcggcgccg tcgtccgcga cccacacctt gccgatgtcg
     6361 agcccgacgc gcgtgaggaa gagttcttgc agctcggtga cccgctcgat gtggcggtcc
     6421 ggatcgacgg tgtggcgcgt ggcggggtag tcggcgaacg cggcggcgag ggtgcgtacg
     6481 gccctgggga cgtcgtcgcg ggtggcgagg cgcaccgtgg gcttgtactc ggtcatggta
     6541 agctccggtg gatccccctg gggagagagg tcggtgattc ggtcaacgag ggagccgact
     6601 gccgacgtgc gctccggagg cttgcagaat gcggaacacc gcgcgggcag gaacagggcc
     6661 cacactaccg ccccacaccc cgcctcccgc accgcccctt cccggccgct gctctcggcg
     6721 cgccctgctg agcagccgct attggccaca gcccatcgcg gtcggcgcgc tgccattgct
     6781 ccctggcgct gtccgtctgc gagggtacta gtgagacgtg cggcttccgt ttgtcacgtc
     6841 cggcacgccg cgaaccgcaa ggaaccttcc cgacttaggg gcggagcagg aagcgtcgcc
     6901 ggggggccca caagggtagc ggcgaagatc cgggtgacgc tgcgaacgga cgtgaagaat
     6961 gtgcgagacc cagggtcggc gccgctgcgt ttcccggaac cacgcccaga gcagccgcgt
     7021 ccctgcgcaa acccagggct gccttggaaa aggcgcaacc ccaaacgctg cgcgtaacca
     7081 ccacacccgc cgcgcttaat gcgccgctac agggcgcgtg gggatacccc ctagagcccc
     7141 agctggttct ttccgcctca gaagccatag agcccaccgc atccccagca tgcctgctat
     7201 tgtcttccca atcctccccc ttgctgtcct gccccacccc accccccaga atagaatgac
     7261 acctactcag acaatgcgat gcaatttcct cat
//
