LOCUS       sequence-173679-        6119 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressed N-term tagged UnaG on Sec61B.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6119)
  AUTHORS   Park JS, Nam E, Lee HK, Lim MH, Rhee HW
  TITLE     In Cellulo Mapping of Subcellular Localized Bilirubin.
  JOURNAL   ACS Chem Biol. 2016 May 27.
  PUBMED    27232847
REFERENCE   2  (bases 1 to 6119)
  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: "ACS Chem 
            Biol. 2016 May 27."
COMMENT     Sequence Label: pCDNA3_UnaG-Flag-Sec61B
FEATURES             Location/Qualifiers
     source          1..6119
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        85..464
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        465..668
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     619..639
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     713..732
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        713..731
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      751..760
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             757..1173
                     /codon_start=1
                     /product="bilirubin-binding green fluorescent protein from 
                     Anguilla japonica (Kumagai et al., 2013)"
                     /label=UnaG
                     /translation="MVEKFVGTWKIADSHNFGEYLKAIGAPKELSDGGDATTPTLYISQ
                     KDGDKMTVKIENGPPTFLDTQVKFKLGEEFDEFPSDRRKGVKSVVNLVGEKLVYVQKWD
                     GKETTYVREIKDGKLVVTLTMGDVVAVRSYRRATE"
     CDS             1174..1197
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1204..1494
                     /codon_start=1
                     /gene="SEC61B"
                     /product="human Sec61-beta subunit of the translocon in the
                     endoplasmic reticulum"
                     /label=Sec61-beta
                     /translation="MPGPTPSGTNVGSSGRSPSKAVAARAAGSTVRQRKNASCGTRSAG
                     RTTSAGTGGMWRFYTEDSPGLKVGPVPVLVMSLLFIASVFMLHIWGKYTRS"
     promoter        complement(1522..1540)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1523..1540)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1560..1577)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1566..1790
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1836..2264
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1923..1942)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2133..2154
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2273..2293)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2278..2607
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2458..2593
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2520..2539
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2674..3468
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2728..2747)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3338..3357
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3642..3763
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3679..3698)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3733..3752
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3812..3828)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3812..3828)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3825..3847)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3836..3852
                     /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(3860..3866,3867..3884,3885..3890))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3905..3926
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4043..4060)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4214..4802)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4294..4313)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4973..5764,5765..5833))
                     /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     5596..5615
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5834..5938)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6013..6032)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gcttgaccga caattgcatg aagaatctgc ttagggttag gcgttttgcg ctgcttcgcg
       61 atgtacgggc cagatatacg cgttgacatt gattattgac tagttattaa tagtaatcaa
      121 ttacggggtc attagttcat agcccatata tggagttccg cgttacataa cttacggtaa
      181 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg
      241 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt
      301 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg
      361 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc
      421 ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc
      481 agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca
      541 ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta
      601 acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa
      661 gcagagctct ctggctaact agagaaccca ctgcttactg gcttatcgaa attaatacga
      721 ctcactatag ggagacccaa gcttgcggcc gccaccatgg ttgaaaaatt tgttggtacg
      781 tggaaaatag cggattctca taattttggc gaatatctga aggctatcgg ggcgcctaaa
      841 gagctgagtg atggaggcga tgccacgact ccaactctgt atatttcaca gaaggacggt
      901 gataaaatga ccgttaagat cgagaatggc ccgcccacct tcctggatac acaggtaaag
      961 tttaaacttg gtgaagaatt tgatgagttc ccgagcgaca gacgcaaagg tgttaaatcc
     1021 gtagtcaact tggtcggaga aaaactcgta tatgtccaaa aatgggatgg caaagagaca
     1081 acctacgtgc gtgaaattaa agacgggaaa ctggttgtga cgctgacaat gggcgacgtg
     1141 gtggcagtgc gctcgtaccg gcgtgcaacc gaagactaca aggatgacga cgataaggaa
     1201 ttcatgcctg gtccgacccc cagtggcact aacgtgggat cctcagggcg ctctcccagc
     1261 aaagcagtgg ccgcccgggc ggcgggatcc actgtccggc agaggaaaaa tgccagctgt
     1321 gggacaagga gtgcaggccg cacaacctcg gcaggcaccg gggggatgtg gcgattctac
     1381 acagaagatt cacctgggct caaagttggc cctgttccag tattggttat gagtcttctg
     1441 ttcatcgctt ctgtatttat gttgcacatt tggggcaagt acactcgttc gtgactcgag
     1501 catgcatcta gagggcccta ttctatagtg tcacctaaat gctagagctc gctgatcagc
     1561 ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt
     1621 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca
     1681 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga
     1741 ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc
     1801 ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag
     1861 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     1921 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     1981 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     2041 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     2101 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     2161 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
     2221 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg
     2281 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     2341 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
     2401 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     2461 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     2521 atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc
     2581 ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga
     2641 tctgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca
     2701 ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc
     2761 ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc
     2821 aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg
     2881 ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg
     2941 gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct
     3001 gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct
     3061 acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa
     3121 gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa
     3181 ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc
     3241 gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt
     3301 ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct
     3361 gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc
     3421 gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg
     3481 ggttcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg
     3541 ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc
     3601 tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt
     3661 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     3721 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt
     3781 cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     3841 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     3901 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     3961 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     4021 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     4081 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     4141 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     4201 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     4261 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     4321 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     4381 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     4441 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     4501 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     4561 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     4621 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     4681 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     4741 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     4801 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     4861 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     4921 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     4981 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     5041 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     5101 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     5161 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     5221 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     5281 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     5341 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     5401 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     5461 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     5521 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     5581 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     5641 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     5701 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     5761 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     5821 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     5881 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     5941 catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg
     6001 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc
     6061 ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaag
//
