LOCUS       sequence-352609-        7439 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Firefly luciferase reporter fused to a prolactin signal sequence for
            ER targeting and KDEL for ER retention and fused to eGFP at the 
            C-terminus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7439)
  AUTHORS   Cortes Sanchon A, Santhosh Kumar H, Mantovani M, Osinnii I, Mateos 
            JM, Kaech A, Shcherbakov D, Akbergenov R, Bottger EC
  TITLE     ER-misfolded proteins become sequestered with mitochondria and 
            impair mitochondrial function.
  JOURNAL   Commun Biol. 2021 Dec 2;4(1):1350. doi: 10.1038/s42003-021-02873-w.
  PUBMED    34857875
REFERENCE   2  (bases 1 to 7439)
  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: "Commun 
            Biol."; date: "2021-12-2"; volume: "4(1):1350. doi"; pages: " 
            10.1038/s42003-021-02873-w"
COMMENT     Sequence Label: pPLN-hFluc-WT-eGFP-KDEL
FEATURES             Location/Qualifiers
     source          1..7439
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             98..1135
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             1114..1122
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     polyA_signal    1159..1207
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(1363..1380)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1534..2122)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1614..1633)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2322..3113,3114..3182))
                     /codon_start=1
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     polyA_signal    3287..3335
                     /note="synthetic polyadenylation signal"
     misc_feature    3349..3440
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     3389..3408
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     enhancer        3591..3970
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3971..4174
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4125..4145
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4171..4195
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             4367..6013
                     /codon_start=1
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="synthetic luc2 version of the luciferase gene"
                     /translation="EDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDAH
                     IEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAPA
                     NDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQSM
                     YTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHAR
                     DPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYKI
                     QSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGYG
                     LTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMSG
                     YVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESILL
                     QHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVVF
                     VDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     CDS             join(6050..6052,6053..6055,6056..6766)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(6050..6052,6053..6055,6056..6766)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(6095..6116)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(6356..6375)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6703..6724
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    6834..6955
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6845..6864)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6899..6918
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        join(7149..7439,1..67)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        7149..7340
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(7172..7192)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      join(7357..7439,1..53)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7419..7438
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
ORIGIN
        1 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
       61 ttgcaaaaag ctcgattctt ctgacactag cgccaccatg aagaagcccg aactcaccgc
      121 taccagcgtt gaaaaatttc tcatcgagaa gttcgacagt gtgagcgacc tgatgcagtt
      181 gtcggagggc gaagagagcc gagccttcag cttcgatgtc ggcggacgcg gctatgtact
      241 gcgggtgaat agctgcgctg atggcttcta caaagaccgc tacgtgtacc gccacttcgc
      301 cagcgctgca ctacccatcc ccgaagtgtt ggacatcggc gagttcagcg agagcctgac
      361 atactgcatc agtagacgcg cccaaggcgt tactctccaa gacctccccg aaacagagct
      421 gcctgctgtg ttacagcctg tcgccgaagc tatggatgct attgccgccg ccgacctcag
      481 tcaaaccagc ggcttcggcc cattcgggcc ccaaggcatc ggccagtaca caacctggcg
      541 ggatttcatt tgcgccattg ctgatcccca tgtctaccac tggcagaccg tgatggacga
      601 caccgtgtcc gccagcgtag ctcaagccct ggacgaactg atgctgtggg ccgaagactg
      661 tcccgaggtg cgccacctcg tccatgccga cttcggcagc aacaacgtcc tgaccgacaa
      721 cggccgcatc accgccgtaa tcgactggtc cgaagctatg ttcggggaca gtcagtacga
      781 ggtggccaac atcttcttct ggcggccctg gctggcttgc atggagcagc agactcgcta
      841 cttcgagcgc cggcatcccg agctggccgg cagccctcgt ctgcgagcct acatgctgcg
      901 catcggcctg gatcagctct accagagcct cgtggacggc aacttcgacg atgctgcctg
      961 ggctcaaggc cgctgcgatg ccatcgtccg cagcggggcc ggcaccgtcg gtcgcacaca
     1021 aatcgctcgc cggagcgcag ccgtatggac cgacggctgc gtcgaggtgc tggccgacag
     1081 cggcaaccgc cggcccagta cacgaccgcg cgctaaggag gtaggtcgag tttaaactct
     1141 agaaccggtc atggccgcaa taaaatatct ttattttcat tacatctgtg tgttggtttt
     1201 ttgtgtgttc gaactagatg ctgtcgaccg atgcccttga gagccttcaa cccagtcagc
     1261 tccttccggt gggcgcgggg catgactatc gtcgccgcac ttatgactgt cttctttatc
     1321 atgcaactcg taggacaggt gccggcagcg ctcttccgct tcctcgctca ctgactcgct
     1381 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     1441 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     1501 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     1561 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     1621 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     1681 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     1741 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     1801 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     1861 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     1921 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     1981 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     2041 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     2101 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     2161 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     2221 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     2281 ttggtctgac agcggccgca aatgctaaac cactgcagtg gttaccagtg cttgatcagt
     2341 gaggcaccga tctcagcgat ctgcctattt cgttcgtcca tagtggcctg actccccgtc
     2401 gtgtagatca ctacgattcg tgagggctta ccatcaggcc ccagcgcagc aatgatgccg
     2461 cgagagccgc gttcaccggc ccccgatttg tcagcaatga accagccagc agggagggcc
     2521 gagcgaagaa gtggtcctgc tactttgtcc gcctccatcc agtctatgag ctgctgtcgt
     2581 gatgctagag taagaagttc gccagtgagt agtttccgaa gagttgtggc cattgctact
     2641 ggcatcgtgg tatcacgctc gtcgttcggt atggcttcgt tcaactctgg ttcccagcgg
     2701 tcaagccggg tcacatgatc acccatatta tgaagaaatg cagtcagctc cttagggcct
     2761 ccgatcgttg tcagaagtaa gttggccgcg gtgttgtcgc tcatggtaat ggcagcacta
     2821 cacaattctc ttaccgtcat gccatccgta agatgctttt ccgtgaccgg cgagtactca
     2881 accaagtcgt tttgtgagta gtgtatacgg cgaccaagct gctcttgccc ggcgtctata
     2941 cgggacaaca ccgcgccaca tagcagtact ttgaaagtgc tcatcatcgg gaatcgttct
     3001 tcggggcgga aagactcaag gatcttgccg ctattgagat ccagttcgat atagcccact
     3061 cttgcaccca gttgatcttc agcatctttt actttcacca gcgtttcggg gtgtgcaaaa
     3121 acaggcaagc aaaatgccgc aaagaaggga atgagtgcga cacgaaaatg ttggatgctc
     3181 atactcgtcc tttttcaata ttattgaagc atttatcagg gttactagta cgtctctcaa
     3241 ggataagtaa gtaatattaa ggtacgggag gtattggaca ggccgcaata aaatatcttt
     3301 attttcatta catctgtgtg ttggtttttt gtgtgaatcg atagtactaa catacgctct
     3361 ccatcaaaac aaaacgaaac aaaacaaact agcaaaatag gctgtcccca gtgcaagtgc
     3421 aggtgccaga acatttctct ggcctaactg gcctcaatat tggccattag ccatattatt
     3481 cattggttat atagcataaa tcaatattgg ctattggcca ttgcatacgt tgtatctata
     3541 tcataatatg tacatttata ttggctcatg tccaatatga ccgccatgtt ggcattgatt
     3601 attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga
     3661 gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg
     3721 cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg
     3781 acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca
     3841 tatgccaagt ccgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc
     3901 ccagtacatg accttacggg actttcctac ttggcagtac atctacgtat tagtcatcgc
     3961 tattaccatg gtgatgcggt tttggcagta caccaatggg cgtggatagc ggtttgactc
     4021 acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa
     4081 tcaacgggac tttccaaaat gtcgtaataa ccccgccccg ttgacgcaaa tgggcggtag
     4141 gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc agatcactag
     4201 aagcttggca acagtaccgt gatacaacca cactcgagcc atgaacatca aaggatcgcc
     4261 atggaaaggg tccctcctgc tgctgctggt gtcaaacctg ctcctgtgcc agagcgtggc
     4321 cccctggaat tcagcttggc aatccggtac tgttggtaaa gccaccgaag atgccaaaaa
     4381 cattaagaag ggcccagcgc cattctaccc actcgaagac gggaccgccg gcgagcagct
     4441 gcacaaagcc atgaagcgct acgccctggt gcccggcacc atcgccttta ccgacgcaca
     4501 tatcgaggtg gacattacct acgccgagta cttcgagatg agcgttcggc tggcagaagc
     4561 tatgaagcgc tatgggctga atacaaacca tcggatcgtg gtgtgcagcg agaatagctt
     4621 gcagttcttc atgcccgtgt tgggtgccct gttcatcggt gtggctgtgg ccccagctaa
     4681 cgacatctac aacgagcgcg agctgctgaa cagcatgggc atcagccagc ccaccgtcgt
     4741 attcgtgagc aagaaagggc tgcaaaagat cctcaacgtg caaaagaagc taccgatcat
     4801 acaaaagatc atcatcatgg atagcaagac cgactaccag ggcttccaaa gcatgtacac
     4861 cttcgtgact tcccatttgc cacccggctt caacgagtac gacttcgtgc ccgagagctt
     4921 cgaccgggac aaaaccatcg ccctgatcat gaacagtagt ggcagtaccg gattgcccaa
     4981 gggcgtagcc ctaccgcacc gcaccgcttg tgtccgattc agtcatgccc gcgaccccat
     5041 cttcggcaac cagatcatcc ccgacaccgc tatcctcagc gtggtgccat ttcaccacgg
     5101 cttcggcatg ttcaccacgc tgggctactt gatctgcggc tttcgggtcg tgctcatgta
     5161 ccgcttcgag gaggagctat tcttgcgcag cttgcaagac tataagattc aatctgccct
     5221 gctggtgccc acactattta gcttcttcgc taagagcact ctcatcgaca agtacgacct
     5281 aagcaacttg cacgagatcg ccagcggcgg ggcgccgctc agcaaggagg taggtgaggc
     5341 cgtggccaaa cgcttccacc taccaggcat ccgccagggc tacggcctga cagaaacaac
     5401 cagcgccatt ctgatcaccc ccgaagggga cgacaagcct ggcgcagtag gcaaggtggt
     5461 gcccttcttc gaggctaagg tggtggactt ggacaccggt aagacactgg gtgtgaacca
     5521 gcgcggcgag ctgtgcgtcc gtggccccat gatcatgagc ggctacgtta acaaccccga
     5581 ggctacaaac gctctcatcg acaaggacgg ctggctgcac agcggcgaca tcgcctactg
     5641 ggacgaggac gagcacttct tcatcgtgga ccggctgaag agcctgatca aatacaaggg
     5701 ctaccaggta gccccagccg aactggagag catcctgctg caacacccca acatcttcga
     5761 cgccggggtc gccggcctgc ccgacgacga tgccggcgag ctgcccgccg cagtcgtcgt
     5821 gctggaacac ggtaaaacca tgaccgagaa ggagatcgtg gactatgtgg ccagccaggt
     5881 tacaaccgcc aagaagctgc gcggtggtgt tgtgttcgtg gacgaggtgc ctaaaggact
     5941 gaccggcaag ttggacgccc gcaagatccg cgagattctc attaaggcca agaagggcgg
     6001 caagatcgcc gtgatccccg ggatcggtgg tggttctgga ggcggttcta tggtgagcaa
     6061 gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa
     6121 cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac
     6181 cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac
     6241 cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt
     6301 cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga
     6361 cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat
     6421 cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta
     6481 caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt
     6541 gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca
     6601 gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac
     6661 ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt
     6721 cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagaaaa agaccaaaga
     6781 tgaactctaa taattctaga gtcggggcgg ccggccgctt cgagcagaca tgataagata
     6841 cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga
     6901 aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa
     6961 caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag
     7021 caagtaaaac ctctacaaat gtggtaaaat cgataaggat ccgtttgcgt attgggcgct
     7081 cttccgctga tctgcgcagc accatggcct gaaataacct ctgaaagagg aacttggtta
     7141 gctaccttct gaggcggaaa gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag
     7201 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     7261 aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     7321 tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
     7381 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggc
//
