LOCUS       sequence-218610-        6306 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses BCCP-PupE in mammalian cell.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6306)
  AUTHORS   Liu Q, Zheng J, Sun W, Huo Y, Zhang L, Hao P, Wang H, Zhuang M
  TITLE     A proximity-tagging system to identify membrane protein-protein 
            interactions.
  JOURNAL   Nat Methods. 2018 Aug 13. pii: 10.1038/s41592-018-0100-5. doi: 
            10.1038/s41592-018-0100-5.
  PUBMED    30104635
REFERENCE   2  (bases 1 to 6306)
  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 
            Methods. 2018 Aug 13. pii: 10.1038/s41592-018-0100-5. doi: 
            10.1038/s41592-018-0100-5."
COMMENT     Sequence Label: pEF6a-BCCP-PupE
FEATURES             Location/Qualifiers
     source          1..6306
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..1179
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          232..1170
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1127..1147
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     1196..1215
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1196..1214
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1277..1294
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1559..1582
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1838..1867
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1883..1900
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1923..1940)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1929..2153
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2199..2627
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2286..2305)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2496..2517
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2636..2656)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2641..2970
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2821..2956
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2883..2902
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3018..3065
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3084..3482
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    3640..3761
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3677..3696)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3731..3750
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3810..3826)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3810..3826)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3823..3845)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3834..3850
                     /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(3858..3864,3865..3882,3883..3888))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3903..3924
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4041..4058)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4201..4789)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4281..4300)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4960..5751,5752..5820))
                     /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     5583..5602
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5821..5925)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6000..6019)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ggctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg
       61 ggaggggtcg gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt
      121 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca
      181 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc
      241 gtgtgtggtt cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt
      301 acttccacct ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
      361 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
      421 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
      481 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
      541 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
      601 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
      661 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
      721 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
      781 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
      841 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
      901 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
      961 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     1021 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     1081 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     1141 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gaattagctt ggtactaata
     1201 cgactcacta tagggagacc caagctggct aggtaagctt ggtaccgagc tcggatccgc
     1261 caccatgagg ggcagccacc accatcatca ccacagccag gatccggccg ggaaggcagg
     1321 cgagggagag atccccgcac ccttggccgg cacggtcagc aaaatcctgg tcaaggaagg
     1381 cgacaccgtg aaggctggac agacggtgtt ggtactggag gcgatgaaga tggagacaga
     1441 gatcaatgcc ccgaccgatg ggaaggtgga gaaggtgttg gttaaggaga gggacgccgt
     1501 gcagggcggt cagggactga tcaagatcgg cgactacgac atcccgacaa ccgccagcct
     1561 ggaagttctg ttccaggggc ccggatccat gaacgcgaaa cagacccaga tcatgggtgg
     1621 cggtggtcgt gacgaagaca atgcggaaga ctctgctcag gcgtctggtc aggttcagat
     1681 caataccgaa ggtgttgact ctctgctgga cgaaatcgac ggcctgctcg aaaacaacgc
     1741 ggaggaattc gttcgttctt acgttcagaa aggtggtgaa taagaattct gcagatatcc
     1801 agcacagtgg cggccgctcg agtctagagg gcccttcgaa caaaaactca tctcagaaga
     1861 ggatctgaat atgcataccg gtcatcatca ccatcaccat tgagtttaaa cccgctgatc
     1921 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     1981 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     2041 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     2101 ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga
     2161 ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
     2221 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     2281 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     2341 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     2401 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     2461 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     2521 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     2581 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat
     2641 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     2701 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     2761 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     2821 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     2881 tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga
     2941 ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc
     3001 ggatctgatc agcacgtgtt gacaattaat catcggcata gtatatcggc atagtataat
     3061 acgacaaggt gaggaactaa accatggcca agcctttgtc tcaagaagaa tccaccctca
     3121 ttgaaagagc aacggctaca atcaacagca tccccatctc tgaagactac agcgtcgcca
     3181 gcgcagctct ctctagcgac ggccgcatct tcactggtgt caatgtatat cattttactg
     3241 ggggaccttg tgcagaactc gtggtgctgg gcactgctgc tgctgcggca gctggcaacc
     3301 tgacttgtat cgtcgcgatc ggaaatgaga acaggggcat cttgagcccc tgcggacggt
     3361 gccgacaggt gcttctcgat ctgcatcctg ggatcaaagc catagtgaag gacagtgatg
     3421 gacagccgac ggcagttggg attcgtgaat tgctgccctc tggttatgtg tgggagggct
     3481 aagcacttcg tggccgagga gcaggactga cacgtgctac gagatttcga ttccaccgcc
     3541 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     3601 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat
     3661 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     3721 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg
     3781 acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     3841 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     3901 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     3961 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     4021 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     4081 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     4141 gcaggaaaga acatgtgagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     4201 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     4261 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     4321 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     4381 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     4441 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     4501 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     4561 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     4621 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     4681 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     4741 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     4801 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     4861 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     4921 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     4981 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     5041 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     5101 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     5161 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     5221 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     5281 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     5341 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     5401 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     5461 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     5521 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     5581 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     5641 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     5701 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     5761 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     5821 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     5881 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     5941 agtgccacct gacgtcgacg gatcgggaga tctcccgatc ccctatggtg cactctcagt
     6001 acaatctgct ctgatgccgc atagttaagc cagtatctgc tccctgcttg tgtgttggag
     6061 gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt gaccgacaat
     6121 tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt acgggccaga
     6181 tatacgcgtt gacattgatt attgactagg cttttgcaaa aagctttgca aagatggata
     6241 aagttttaaa cagagaggaa tctttgcagc taatggacct tctaggtctt gaaaggagtg
     6301 ggaatt
//
