LOCUS       sequence-455455-        7016 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BioID experiment.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7016)
  TITLE     Ori Lab colocalization and BioID plasmids 2024
REFERENCE   2  (bases 1 to 7016)
  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"
COMMENT     Sequence Label: pcDNA-pDEST_TMEM192_BirA_Flag
FEATURES             Location/Qualifiers
     source          1..7016
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     585..604
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(823..927)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1002..1021)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1193..1572
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1573..1776
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1727..1747
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    1778..1796
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1799..1817
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     1827..1851
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    1945..1969
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      1988..1997
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(2834..2858)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             join(2869..3219,3220..3222,3223..3831)
                     /codon_start=1
                     /product="promiscuous R118G mutant of the E. coli biotin 
                     protein ligase (Cronan, 2005)"
                     /label=BirA(R118G)
                     /note="also known as BirA* or BioID"
                     /translation="MKDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRD
                     WGVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSG
                     DACIAEYQQAGRGGRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRK
                     LGADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQG
                     WITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIG
                     DKEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     CDS             3862..3885
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(3926..3943)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3932..4156
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4289..4308)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4440..4487
                     /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)."
     CDS             4495..5517
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    5647..5768
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5684..5703)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5738..5757
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5817..5833)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5817..5833)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5830..5852)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5841..5857
                     /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(5865..5871,5872..5889,5890..5895))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5910..5931
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6048..6065)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6219..6807)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6299..6318)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6978..7016,1..753,754..822))
                     /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"
ORIGIN
        1 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
       61 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
      121 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
      181 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
      241 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
      301 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
      361 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
      421 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
      481 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
      541 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
      601 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
      661 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
      721 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
      781 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
      841 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
      901 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcga
      961 cggatcggga gatctcccga tcccctatgg tgcactctca gtacaatctg ctctgatgcc
     1021 gcatagttaa gccagtatct gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg
     1081 agcaaaattt aagctacaac aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt
     1141 agggttaggc gttttgcgct gcttcgcgat gtacgggcca gatatacgcg ttgacattga
     1201 ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg
     1261 gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc
     1321 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat
     1381 tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat
     1441 catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
     1501 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc
     1561 gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac
     1621 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa
     1681 aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt
     1741 aggcgtgtac ggtgggaggt ctatataagc agagctctcc ctatcagtga tagagatctc
     1801 cctatcagtg atagagatcg tcgacgagct cgtttagtga accgtcagat cgcctggaga
     1861 cgccatccac gctgttttga cctccataga agacaccggg accgatccag cctccggact
     1921 ctagcgttta aacttaagct ttagacaagt ttgtacaaaa aagcaggctc cgcggccgcc
     1981 cccttcacca tggcggcggg gggcaggatg gaggacggtt ccttggatat cacccagagt
     2041 attgaagacg acccacttct ggatgcccag cttctcccac accactcatt acaagctcac
     2101 tttagacccc gattccatcc tcttcctaca gtcatcatag tgaatcttct gtggtttatt
     2161 catctcgtgt ttgttgtttt agcattttta acaggtgtgc tttgttctta tcctaatcca
     2221 aatgaggaca agtgcccagg aaattacaca aacccattga aagttcagac ggttataatc
     2281 cttgggaaag ttattttgtg gattctccat ttactccttg aatgctacat ccagtatcac
     2341 cacagcaaaa tcagaaaccg aggctataac ttgatctacc gatcaacaag gcatctcaag
     2401 agacttgcgt tgatgataca gtcctctggc aacacagtgc ttctcctcat actgtgcatg
     2461 cagcactcct tcccagagcc tggcagattg tatcttgacc tcattctggc catcttggca
     2521 ctggaactca tctgttccct gatatgtctc ctcatttaca cagtgaaaat ccggagattt
     2581 aataaagcta aaccagagcc tgatatactt gaagaagaaa aaatctatgc ttaccccagc
     2641 aatattacct cggagactgg attcagaact atttcaagcc tagaagaaat tgttgaaaag
     2701 caaggagaca ccattgaata cctgaagcga cacaatgcgc tgctgagtaa gcgattgttg
     2761 gctctcactt cctcagacct gggctgtcag ccaagtagaa cgggaggtgg aggtagtaag
     2821 ggtgggcgcg ccgacccagc tttcttgtac aaagtggttg attctagaat gaaggacaac
     2881 accgtgcccc tgaagctgat cgccctgctg gccaacggcg agttccacag cggcgagcag
     2941 ctgggcgaga ccctgggcat gagccgggcc gccatcaaca agcacatcca gaccctgcgg
     3001 gactggggcg tggacgtgtt caccgtgccc ggcaagggct acagcctgcc cgagcccatc
     3061 cagctgctga acgccaagca gatcctgggc cagctggacg gcggcagcgt ggccgtgctg
     3121 cccgtgatcg acagcaccaa ccagtacctg ctggaccgga tcggcgagct gaagagcggc
     3181 gacgcctgca tcgccgagta ccagcaggcc ggccggggcg gccggggccg gaagtggttc
     3241 agccccttcg gcgccaacct gtacctgagc atgttctggc ggctggagca gggccccgcc
     3301 gccgccatcg gcctgagcct ggtgatcggc atcgtgatgg ccgaggtgct gcggaagctg
     3361 ggcgccgaca aggtgcgggt gaagtggccc aacgacctgt acctgcagga ccggaagctg
     3421 gccggcatcc tggtggagct gaccggcaag accggcgacg ccgcccagat cgtgatcggc
     3481 gccggcatca acatggccat gcggcgggtg gaggagagcg tggtgaacca gggctggatc
     3541 accctgcagg aggccggcat caacctggac cggaacaccc tggccgccat gctgatccgg
     3601 gagctgcggg ccgccctgga gctgttcgag caggagggcc tggcccccta cctgagccgg
     3661 tgggagaagc tggacaactt catcaaccgg cccgtgaagc tgatcatcgg cgacaaggag
     3721 atcttcggca tcagccgggg catcgacaag cagggcgccc tgctgctgga gcaggacggc
     3781 atcatcaagc cctggatggg cggcgagatc agcctgcgga gcgccgagaa ggggggatct
     3841 ggccccggcg gcggcgcgcc agactacaaa gacgatgacg ataaataact cgagtctaga
     3901 gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     3961 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     4021 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     4081 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat
     4141 gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc
     4201 cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     4261 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     4321 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     4381 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtacctag
     4441 aagttcctat tccgaagttc ctattctcta gaaagtatag gaacttcctt ggccaaaaag
     4501 cctgaactca ccgcgacgtc tgtcgagaag tttctgatcg aaaagttcga cagcgtctcc
     4561 gacctgatgc agctctcgga gggcgaagaa tctcgtgctt tcagcttcga tgtaggaggg
     4621 cgtggatatg tcctgcgggt aaatagctgc gccgatggtt tctacaaaga tcgttatgtt
     4681 tatcggcact ttgcatcggc cgcgctcccg attccggaag tgcttgacat tggggagttc
     4741 agcgagagcc tgacctattg catctcccgc cgtgcacagg gtgtcacgtt gcaagacctg
     4801 cctgaaaccg aactgcccgc tgttctgcag ccggtcgcgg aggccatgga tgcgatcgct
     4861 gcggccgatc ttagccagac gagcgggttc ggcccattcg gaccgcaagg aatcggtcaa
     4921 tacactacat ggcgtgattt catatgcgcg attgctgatc cccatgtgta tcactggcaa
     4981 actgtgatgg acgacaccgt cagtgcgtcc gtcgcgcagg ctctcgatga gctgatgctt
     5041 tgggccgagg actgccccga agtccggcac ctcgtgcacg cggatttcgg ctccaacaat
     5101 gtcctgacgg acaatggccg cataacagcg gtcattgact ggagcgaggc gatgttcggg
     5161 gattcccaat acgaggtcgc caacatcttc ttctggaggc cgtggttggc ttgtatggag
     5221 cagcagacgc gctacttcga gcggaggcat ccggagcttg caggatcgcc gcggctccgg
     5281 gcgtatatgc tccgcattgg tcttgaccaa ctctatcaga gcttggttga cggcaatttc
     5341 gatgatgcag cttgggcgca gggtcgatgc gacgcaatcg tccgatccgg agccgggact
     5401 gtcgggcgta cacaaatcgc ccgcagaagc gcggccgtct ggaccgatgg ctgtgtagaa
     5461 gtactcgccg atagtggaaa ccgacgcccc agcactcgtc cgagggcaaa ggaatagcac
     5521 gtactacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt
     5581 ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc
     5641 caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     5701 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     5761 gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca
     5821 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     5881 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     5941 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     6001 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     6061 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     6121 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     6181 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     6241 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     6301 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     6361 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     6421 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     6481 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     6541 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     6601 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     6661 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     6721 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     6781 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     6841 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     6901 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     6961 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgat
//
