LOCUS       sequence-435399-        7565 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expressed the NLS-Halo-eGFP-BASU fusion protein via stable Flp-In 
            integration.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7565)
  AUTHORS   Hanswillemenke A, Hofacker DT, Sorgenfrei M, Fruhner C, 
            Franz-Wachtel M, Schwarzer D, Macek B, Stafforst T
  TITLE     Profiling the interactome of oligonucleotide drugs by proximity 
            biotinylation.
  JOURNAL   Nat Chem Biol. 2024 Jan 17. doi: 10.1038/s41589-023-01530-z.
  PUBMED    38233583
REFERENCE   2  (bases 1 to 7565)
  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"; journalName: "Nat Chem 
            Biol. 2024 Jan 17. doi: 10.1038/s41589-023-01530-z."
COMMENT     Sequence Label: pcDNA5_NLS-Halo-eGFP-BASU
FEATURES             Location/Qualifiers
     source          1..7565
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(299..319)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(442..466)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    476..494
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    497..515
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        complement(517..720)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(546..566)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        721..1100
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     1272..1291
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        1366..1470
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1471..1539,1540..2331)
                     /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(1689..1708)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2502..3090
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2991..3010
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3244..3261
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3378..3399
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3414..3419,3420..3437,3438..3444)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3452..3468
                     /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     3457..3479
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3476..3492
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3476..3492
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    3541..3662
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3552..3571)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3606..3625
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(3792..4814)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             complement(3792..4814)
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     protein_bind    complement(4822..4869)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     primer_bind     5001..5020
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    5153..5377
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     5366..5383
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(join(6217..6927,6928..6930))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             complement(join(6217..6927,6928..6930))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(6259..6280)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     6608..6627
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6867..6888
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
ORIGIN
        1 ccagggcttc gatgaaggca tccatgaagc ggacgtggtc gtcgaagaaa taacccaggt
       61 ctggtttgtc ggatttgccc ataccgatca ggtctggagc aatgcagcga tgggtcggtg
      121 caacatgcgg gatgatgttg cgccacacgt aggaggaggt cgggttaccg tgcaggaaca
      181 gcacaggggt gccatcgcgc ggaccaacat cgacgtagtg catgcgctcg cccaggactt
      241 ccacataatg ggggtcgaat ggaaagccag taccgatttc tgcacctgag cctggagcaa
      301 cttttctttt cttttttggt cccatgcggc cgcggatccg agctcggtac caagcttaag
      361 tttaaacgct agagtccgga ggctggatcg gtcccggtgt cttctatgga ggtcaaaaca
      421 gcgtggatgg cgtctccagg cgatctgacg gttcactaaa cgagctcgtc gacgatctct
      481 atcactgata gggagatctc tatcactgat agggagagct ctgcttatat agacctccca
      541 ccgtacacgc ctaccgccca tttgcgtcaa tggggcggag ttgttacgac attttggaaa
      601 gtcccgttga ttttggtgcc aaaacaaact cccattgacg tcaatggggt ggagacttgg
      661 aaatccccgt gagtcaaacc gctatccacg cccattgatg tactgccaaa accgcatcac
      721 catggtaata gcgatgacta atacgtagat gtactgccaa gtaggaaagt cccataaggt
      781 catgtactgg gcataatgcc aggcgggcca tttaccgtca ttgacgtcaa tagggggcgt
      841 acttggcata tgatacactt gatgtactgc caagtgggca gtttaccgta aatactccac
      901 ccattgacgt caatggaaag tccctattgg cgttactatg ggaacatacg tcattattga
      961 cgtcaatggg cgggggtcgt tgggcggtca gccaggcggg ccatttaccg taagttatgt
     1021 aacgcggaac tccatatatg ggctatgaac taatgacccc gtaattgatt actattaata
     1081 actagtcaat aatcaatgtc aacgcgtata tctggcccgt acatcgcgaa gcagcgcaaa
     1141 acgcctaacc ctaagcagat tcttcatgca attgtcggtc aagccttgcc ttgttgtagc
     1201 ttaaattttg ctcgcgcact actcagcgac ctccaacaca caagcaggga gcagatactg
     1261 gcttaactat gcggcatcag agcagattgt actgagagtg caccataggg gatcgggaga
     1321 tctcccgatc cgtcgacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     1381 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     1441 atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt
     1501 attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa
     1561 gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac
     1621 agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt
     1681 aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt
     1741 cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat
     1801 cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac
     1861 actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg
     1921 cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc
     1981 ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa
     2041 ctattaactg gcgaactact tactctagct tcccggcaac aattaataga ctggatggag
     2101 gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct
     2161 gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat
     2221 ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa
     2281 cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac
     2341 caagtttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
     2401 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     2461 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     2521 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     2581 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     2641 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     2701 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     2761 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     2821 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     2881 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     2941 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     3001 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     3061 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     3121 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     3181 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
     3241 cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc
     3301 gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc
     3361 agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac
     3421 tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga
     3481 aacagctatg accatgatta cgccaagctc tagctagagg tcgacggtat acagacatga
     3541 taagatacat tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta
     3601 tttgtgaaat ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag
     3661 ttggggtggg cgaagaactc cagcatgaga tccccgcgct ggaggatcat ccagccggcg
     3721 tcccggaaaa cgattccgaa gcccaacctt tcatagaagg cggcggtgga atcgaaatct
     3781 cgtagtacgt gctattcctt tgccctcgga cgagtgctgg ggcgtcggtt tccactatcg
     3841 gcgagtactt ctacacagcc atcggtccag acggccgcgc ttctgcgggc gatttgtgta
     3901 cgcccgacag tcccggctcc ggatcggacg attgcgtcgc atcgaccctg cgcccaagct
     3961 gcatcatcga aattgccgtc aaccaagctc tgatagagtt ggtcaagacc aatgcggagc
     4021 atatacgccc ggagccgcgg cgatcctgca agctccggat gcctccgctc gaagtagcgc
     4081 gtctgctgct ccatacaagc caaccacggc ctccagaaga agatgttggc gacctcgtat
     4141 tgggaatccc cgaacatcgc ctcgctccag tcaatgaccg ctgttatgcg gccattgtcc
     4201 gtcaggacat tgttggagcc gaaatccgcg tgcacgaggt gccggacttc ggggcagtcc
     4261 tcggcccaaa gcatcagctc atcgagagcc tgcgcgacgg acgcactgac ggtgtcgtcc
     4321 atcacagttt gccagtgata cacatgggga tcagcaatcg cgcatatgaa atcacgccat
     4381 gtagtgtatt gaccgattcc ttgcggtccg aatgggccga acccgctcgt ctggctaaga
     4441 tcggccgcag cgatcgcatc catggcctcc gcgaccggct gcagaacagc gggcagttcg
     4501 gtttcaggca ggtcttgcaa cgtgacaccc tgtgcacggc gggagatgca ataggtcagg
     4561 ctctcgctga attccccaat gtcaagcact tccggaatcg ggagcgcggc cgatgcaaag
     4621 tgccgataaa cataacgatc tttgtagaaa ccatcggcgc agctatttac ccgcaggaca
     4681 tatccacgcc ctcctacatc gaagctgaaa gcacgagatt cttcgccctc cgagagctgc
     4741 atcaggtcgg agacgctgtc gaacttttcg atcagaaact tctcgacaga cgtcgcggtg
     4801 agttcaggct ttttggccaa ggaagttcct atactttcta gagaatagga acttcggaat
     4861 aggaacttct aggtacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt
     4921 aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg
     4981 gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca
     5041 agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag
     5101 ggcgcgtggg gataccccct agagccccag ctggttcttt ccgcctcaga agccatagag
     5161 cccaccgcat ccccagcatg cctgctattg tcttcccaat cctccccctt gctgtcctgc
     5221 cccaccccac cccccagaat agaatgacac ctactcagac aatgcgatgc aatttcctca
     5281 ttttattagg aaaggacagt gggagtggca ccttccaggg tcaaggaagg cacgggggag
     5341 gggcaaacaa cagatggctg gcaactagaa ggcacagtcg aggctgatca gcgggtttaa
     5401 acgggcccat cgattctaga attagcgcag gctgatgtcg gcgctgtaga ttttcttgat
     5461 gccctcgttt gtttccagca gcagaacgcc ctcgtcatca attcccaggg ccttgccgta
     5521 gaaggtgccg ttcagtgtgc gggcgcgcat gttggtgccg attccgaggg cgtagctctc
     5581 ccacagcagt ttgatagggg tgaagccgtg ggtcatgtag tcgcggtacc gcttctcgaa
     5641 gcacagcagg atgtgctgga tcacgccagc gcgatcaatc ttctcgccag cggcctgaga
     5701 caggcttgtg gcgatatcct tcagctcgtc ggggaagtcg ttaggctgct ggttcacatt
     5761 gatcccgatg ccgatgatca cgctgcggac tctgtcctct tcggcctgca tctcggtcag
     5821 gatgcccacg gttttcttgc cgttgatcag gatgtcgttg ggccacttga tgtcggtctg
     5881 gattccggcg gcttcttcaa ttccctgcac cacagccaca gcagccagca gtgtcagctg
     5941 aggggttttc tggagaggga tgtcagggcg caggatcagg ctcatccaga cgccgttgcc
     6001 ttcttggctg tgccacacgc ggctcattcc gccgcggcca gctgtctgct tatcggccac
     6061 caccagtgtt ccctcagggg cattgttgtt ggccagctcg tgggctgttt tctgggtgct
     6121 gctcagcacg tcgtggtaga tcaggtgctg gcccatcact tcggttttca ggccgaagcg
     6181 gatctcgctc tcgctaagtt ttccgccggt gaccggcttg tacagctcgt ccatgccgag
     6241 agtgatcccg gcggcggtca cgaactccag caggaccatg tgatcgcgct tctcgttggg
     6301 gtctttgctc agggcggact gggtgctcag gtagtggttg tcgggcagca gcacggggcc
     6361 gtcgccgatg ggggtgttct gctggtagtg gtcggcgagc tgcacgctgc cgtcctcgat
     6421 gttgtggcgg atcttgaagt tcaccttgat gccgttcttc tgcttgtcgg ccatgatata
     6481 gacgttgtgg ctgttgtagt tgtactccag cttgtgcccc aggatgttgc cgtcctcctt
     6541 gaagtcgatg cccttcagct cgatgcggtt caccagggtg tcgccctcga acttcacctc
     6601 ggcgcgggtc ttgtagttgc cgtcgtcctt gaagaagatg gtgcgctcct ggacgtagcc
     6661 ttcgggcatg gcggacttga agaagtcgtg ctgcttcatg tggtcggggt agcggctgaa
     6721 gcactgcacg ccgtaggtca gggtggtcac gagggtgggc cagggcacgg gcagcttgcc
     6781 ggtggtgcag atgaacttca gggtcagctt gccgtaggtg gcatcgccct cgccctcgcc
     6841 ggacacgctg aacttgtggc cgtttacgtc gccgtccagc tcgaccagga tgggcaccac
     6901 cccggtgaac agctcctcgc ccttgctcac gccagaccct ggcgcgcctc cgcctgctgg
     6961 ggttggcttc tcgagcgtcg acagccagcg cgcgatctcg ctgccgatca ggtccgggtt
     7021 gtcttcttgc agcagattca gacccgggcc gatgtccaca gccttgcagt taggcaggct
     7081 tttggccagg cgagcggctt cggccggtgg gatcagaacg cctggggtgc cccagaacag
     7141 cagcttcggg acaggggact ggtgcagcca gtccatgtat tcttcgacca gcgcgacgat
     7201 gttcgctggc tcaccggcga ttggcagctc gtttgggaag cgccacagtg gctcgcggtc
     7261 aacaggattc aggaacggct cgcggtaatg gtccatctcg acttcagtca gcgggcggac
     7321 gacacccatc ggcagcgtac cctcgataaa aacgttctga tcgatgatca gcttgcggcc
     7381 gacgtcggtg gtgcggaagg cctggaaggt ctcgcgggca aattctggcc attcgtccca
     7441 ggtcgggata gggcggatga actccataaa tgcaatacct ttgacgcgct ctggattgcg
     7501 cttggcccag tggaaaccca gagcggagcc ccagtcgtga atgaccagga cgacctcttc
     7561 cagac
//
