LOCUS       sequence-112829-        6793 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible expression of proteins fused to an N-terminal Flag-HA tag 
            (FH).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6793)
  AUTHORS   Beck DB, Narendra V, Drury WJ 3rd, Casey R, Jansen PW, Yuan ZF, 
            Garcia BA, Vermeulen M, Bonasio R
  TITLE     In Vivo Proximity Labeling for the Detection of Protein-Protein and 
            Protein-RNA Interactions.
  JOURNAL   J Proteome Res. 2014 Dec 5;13(12):6135-43. doi: 10.1021/pr500196b. 
            Epub 2014 Oct 29.
  PUBMED    25311790
REFERENCE   2  (bases 1 to 6793)
  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"; doi: "10.1021/pr500196b";
            journalName: "J Proteome Res"; date: "2014-12-5- 5"; volume: "13"; 
            issue: "12"; pages: "6135-43"
COMMENT     Sequence Label: pINTO-N-FH
FEATURES             Location/Qualifiers
     source          1..6793
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(52..71)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1223..1602
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1603..1806
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1757..1777
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    1808..1826
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1829..1847
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     1857..1881
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      1964..1973
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1973..1996
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2006..2032
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2039..2059
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(2180..2197)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2186..2410
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2456..2884
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2543..2562)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2753..2774
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2893..2913)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2898..3227
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3078..3213
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3140..3159
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3275..3322
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3341..3715
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    3845..3966
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3882..3901)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3936..3955
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4015..4031)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4015..4031)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4028..4050)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4039..4055
                     /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(4063..4069,4070..4087,4088..4093))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4108..4129
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4878..4895)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5049..5634)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5129..5148)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5805..6596,6597..6665))
                     /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     6428..6447
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6666..6770)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctgacgtcga cggatcggga gatctcccga tcccctatgg tgcactctca gtacaatctg
       61 ctctgatgcc gcatagttaa gccagtatct gctccctgct tgtgtgttgg aggtcgctga
      121 gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc ttgaccgaca attgttaatt
      181 aagctagcag tctgcaatga gttcgagttg gcgcgcctgt cattctaaat ctctctttca
      241 gcctaaagct ttttccccgt atccccccag gtgtctgcag gctcaaagag cagcgagaag
      301 cgttcagagg aaagcgatcc cgtgccacct tccccgtgcc cgggctgtcc ccgcacgctg
      361 ccggctcggg gatgcggggg gagcgccgga ccggagcgga gccccgggcg gctcgctgct
      421 gccccctagc gggggaggga cgtaattaca tccctggggg ctttgggagg gggctgtccc
      481 cgtgagctcc caggcgcgcc tgtcattcta aatctctctt tcagcctaaa gctttttccc
      541 cgtatccccc caggtgtctg caggctcaaa gagcagcgag aagcgttcag aggaaagcga
      601 tcccgtgcca ccttccccgt gcccgggctg tccccgcacg ctgccggctc ggggatgcgg
      661 ggggagcgcc ggaccggagc ggagccccgg gcggctcgct gctgccccct agcgggggag
      721 ggacgtaatt acatccctgg gggctttggg ggggggctgt ccccgtgagc tcccaggcgc
      781 gccaactcgg tactcgggga tgctagcgca ccatcaacga ggaggacatc ttggctaacc
      841 aacccctgtt gttacagtcc atccaggaca gacttggctc gctggtgggc caggacagcg
      901 ggtatgtggg gggtcttccc aagaacgtta aggaaaagct gctgagcttg aagactttac
      961 aaagcgagct attcgaagtt gaaaaggaat ttcaggtgga gatgtttgag ctagagaaca
     1021 agtttctgca gaagtacaag cccatttggg agcagcggtc caggatcatt tcaggacaag
     1081 agcaacccaa gccggaacag atcgcaaagg gccaagagat tgtggagtca ctcaatgaga
     1141 cggagttgca attgcatgaa gaatctgctt agggttaggc gttttgcgct gcttcgcgat
     1201 gtacgggcca gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt
     1261 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
     1321 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     1381 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     1441 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
     1501 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
     1561 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
     1621 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
     1681 gacgtcaatg ggagtttgtt ttggaaccaa aatcaacggg actttccaaa atgtcgtaac
     1741 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
     1801 agagctctcc ctatcagtga tagagatctc cctatcagtg atagagatcg tcgacgagct
     1861 cgtttagtga accgtcagat cgcctggaga cgccatccac gctgttttga cctccataga
     1921 agacaccggg accgatccag cctccggact ctagcgttta aaggccacca tggattacaa
     1981 ggatgacgac gataagggct ccggataccc atacgatgtg ccagattacg ctggcagcga
     2041 gaacctgtac ttccagggct ccggaaccgg tagcggtacc gagctcggat ccactagtcc
     2101 agtgtggtgg aattctgcag atatccagca cagtggcggc cgctcgagtc tagagggccc
     2161 gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc
     2221 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa
     2281 aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg
     2341 gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg
     2401 ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg
     2461 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     2521 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     2581 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     2641 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     2701 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     2761 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     2821 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     2881 aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag
     2941 gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag
     3001 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc
     3061 cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc
     3121 atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc tctgagctat
     3181 tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcccgggag
     3241 cttgtatatc cattttcgga tctgatcagc acgtgttgac aattaatcat cggcatagta
     3301 tatcggcata gtataatacg acaaggtgag gaactaaacc atggccaagt tgaccagtgc
     3361 cgttccggtg ctcaccgcgc gcgacgtcgc cggagcggtc gagttctgga ccgaccggct
     3421 cgggttctcc cgggacttcg tggaggacga cttcgccggt gtggtccggg acgacgtgac
     3481 cctgttcatc agcgcggtcc aggaccaggt ggtgccggac aacaccctgg cctgggtgtg
     3541 ggtgcgcggc ctggacgagc tgtacgccga gtggtcggag gtcgtgtcca cgaacttccg
     3601 ggacgcctcc gggccggcca tgaccgagat cggcgagcag ccgtgggggc gggagttcgc
     3661 cctgcgcgac ccggccggca actgcgtgca cttcgtggcc gaggagcagg actgacacgt
     3721 gctacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     3781 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     3841 ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     3901 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     3961 atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata
     4021 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     4081 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     4141 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     4201 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gctagtctgc aatgagttcg
     4261 agttggcgcg cctgtcattc taaatctctc tttcagccta aagctttttc cccgtatccc
     4321 cccaggtgtc tgcaggctca aagagcagcg agaagcgttc agaggaaagc gatcccgtgc
     4381 caccttcccc gtgcccgggc tgtccccgca cgctgccggc tcggggatgc ggggggagcg
     4441 ccggaccgga gcggagcccc gggcggctcg ctgctgcccc ctagcggggg agggacgtaa
     4501 ttacatccct gggggctttg ggagggggct gtccccgtga gctcccaggc gcgcctgtca
     4561 ttctaaatct ctctttcagc ctaaagcttt ttccccgtat ccccccaggt gtctgcaggc
     4621 tcaaagagca gcgagaagcg ttcagaggaa agcgatcccg tgccaccttc cccgtgcccg
     4681 ggctgtcccc gcacgctgcc ggctcgggga tgcgggggga gcgccggacc ggagcggagc
     4741 cccgggcggc tcgctgctgc cccctagcgg gggagggacg taattacatc cctgggggct
     4801 ttgggggggg gctgtccccg tgagctccca ggcgcgccaa ctcggtactc ggggatttaa
     4861 ttaagctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     4921 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     4981 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     5041 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     5101 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     5161 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     5221 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     5281 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     5341 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     5401 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     5461 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     5521 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     5581 agcggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     5641 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     5701 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     5761 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     5821 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     5881 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     5941 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     6001 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     6061 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     6121 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     6181 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     6241 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     6301 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     6361 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     6421 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     6481 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     6541 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     6601 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     6661 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     6721 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     6781 cgaaaagtgc cac
//
