LOCUS       sequence-352605-        6686 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Firefly luciferase reporter fused to a prolactin signal sequence for
            ER targeting and KDEL for ER retention.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6686)
  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 6686)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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-KDEL
FEATURES             Location/Qualifiers
     source          1..6686
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     42..61
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             160..1197
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             1176..1184
                     /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    1221..1269
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(1425..1442)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1596..2184)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1676..1695)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2384..3175,3176..3244))
                     /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    3349..3397
                     /note="synthetic polyadenylation signal"
     misc_feature    3411..3502
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     3451..3470
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     enhancer        3653..4032
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4033..4236
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4187..4207
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4233..4257
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             4429..6075
                     /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"
     polyA_signal    6143..6264
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6154..6173)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6208..6227
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        join(6458..6686,1..129)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        6458..6649
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(6481..6501)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      join(6666..6686,1..115)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
       61 gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
      121 ttttgcaaaa agctcgattc ttctgacact agcgccacca tgaagaagcc cgaactcacc
      181 gctaccagcg ttgaaaaatt tctcatcgag aagttcgaca gtgtgagcga cctgatgcag
      241 ttgtcggagg gcgaagagag ccgagccttc agcttcgatg tcggcggacg cggctatgta
      301 ctgcgggtga atagctgcgc tgatggcttc tacaaagacc gctacgtgta ccgccacttc
      361 gccagcgctg cactacccat ccccgaagtg ttggacatcg gcgagttcag cgagagcctg
      421 acatactgca tcagtagacg cgcccaaggc gttactctcc aagacctccc cgaaacagag
      481 ctgcctgctg tgttacagcc tgtcgccgaa gctatggatg ctattgccgc cgccgacctc
      541 agtcaaacca gcggcttcgg cccattcggg ccccaaggca tcggccagta cacaacctgg
      601 cgggatttca tttgcgccat tgctgatccc catgtctacc actggcagac cgtgatggac
      661 gacaccgtgt ccgccagcgt agctcaagcc ctggacgaac tgatgctgtg ggccgaagac
      721 tgtcccgagg tgcgccacct cgtccatgcc gacttcggca gcaacaacgt cctgaccgac
      781 aacggccgca tcaccgccgt aatcgactgg tccgaagcta tgttcgggga cagtcagtac
      841 gaggtggcca acatcttctt ctggcggccc tggctggctt gcatggagca gcagactcgc
      901 tacttcgagc gccggcatcc cgagctggcc ggcagccctc gtctgcgagc ctacatgctg
      961 cgcatcggcc tggatcagct ctaccagagc ctcgtggacg gcaacttcga cgatgctgcc
     1021 tgggctcaag gccgctgcga tgccatcgtc cgcagcgggg ccggcaccgt cggtcgcaca
     1081 caaatcgctc gccggagcgc agccgtatgg accgacggct gcgtcgaggt gctggccgac
     1141 agcggcaacc gccggcccag tacacgaccg cgcgctaagg aggtaggtcg agtttaaact
     1201 ctagaaccgg tcatggccgc aataaaatat ctttattttc attacatctg tgtgttggtt
     1261 ttttgtgtgt tcgaactaga tgctgtcgac cgatgccctt gagagccttc aacccagtca
     1321 gctccttccg gtgggcgcgg ggcatgacta tcgtcgccgc acttatgact gtcttcttta
     1381 tcatgcaact cgtaggacag gtgccggcag cgctcttccg cttcctcgct cactgactcg
     1441 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     1501 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     1561 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     1621 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     1681 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     1741 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     1801 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     1861 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     1921 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     1981 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     2041 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     2101 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     2161 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     2221 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     2281 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     2341 acttggtctg acagcggccg caaatgctaa accactgcag tggttaccag tgcttgatca
     2401 gtgaggcacc gatctcagcg atctgcctat ttcgttcgtc catagtggcc tgactccccg
     2461 tcgtgtagat cactacgatt cgtgagggct taccatcagg ccccagcgca gcaatgatgc
     2521 cgcgagagcc gcgttcaccg gcccccgatt tgtcagcaat gaaccagcca gcagggaggg
     2581 ccgagcgaag aagtggtcct gctactttgt ccgcctccat ccagtctatg agctgctgtc
     2641 gtgatgctag agtaagaagt tcgccagtga gtagtttccg aagagttgtg gccattgcta
     2701 ctggcatcgt ggtatcacgc tcgtcgttcg gtatggcttc gttcaactct ggttcccagc
     2761 ggtcaagccg ggtcacatga tcacccatat tatgaagaaa tgcagtcagc tccttagggc
     2821 ctccgatcgt tgtcagaagt aagttggccg cggtgttgtc gctcatggta atggcagcac
     2881 tacacaattc tcttaccgtc atgccatccg taagatgctt ttccgtgacc ggcgagtact
     2941 caaccaagtc gttttgtgag tagtgtatac ggcgaccaag ctgctcttgc ccggcgtcta
     3001 tacgggacaa caccgcgcca catagcagta ctttgaaagt gctcatcatc gggaatcgtt
     3061 cttcggggcg gaaagactca aggatcttgc cgctattgag atccagttcg atatagccca
     3121 ctcttgcacc cagttgatct tcagcatctt ttactttcac cagcgtttcg gggtgtgcaa
     3181 aaacaggcaa gcaaaatgcc gcaaagaagg gaatgagtgc gacacgaaaa tgttggatgc
     3241 tcatactcgt cctttttcaa tattattgaa gcatttatca gggttactag tacgtctctc
     3301 aaggataagt aagtaatatt aaggtacggg aggtattgga caggccgcaa taaaatatct
     3361 ttattttcat tacatctgtg tgttggtttt ttgtgtgaat cgatagtact aacatacgct
     3421 ctccatcaaa acaaaacgaa acaaaacaaa ctagcaaaat aggctgtccc cagtgcaagt
     3481 gcaggtgcca gaacatttct ctggcctaac tggcctcaat attggccatt agccatatta
     3541 ttcattggtt atatagcata aatcaatatt ggctattggc cattgcatac gttgtatcta
     3601 tatcataata tgtacattta tattggctca tgtccaatat gaccgccatg ttggcattga
     3661 ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg
     3721 gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc
     3781 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat
     3841 tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat
     3901 catatgccaa gtccgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
     3961 gcccagtaca tgaccttacg ggactttcct acttggcagt acatctacgt attagtcatc
     4021 gctattacca tggtgatgcg gttttggcag tacaccaatg ggcgtggata gcggtttgac
     4081 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa
     4141 aatcaacggg actttccaaa atgtcgtaat aaccccgccc cgttgacgca aatgggcggt
     4201 aggcgtgtac ggtgggaggt ctatataagc agagctcgtt tagtgaaccg tcagatcact
     4261 agaagcttgg caacagtacc gtgatacaac cacactcgag ccatgaacat caaaggatcg
     4321 ccatggaaag ggtccctcct gctgctgctg gtgtcaaacc tgctcctgtg ccagagcgtg
     4381 gccccctgga attcagcttg gcaatccggt actgttggta aagccaccga agatgccaaa
     4441 aacattaaga agggcccagc gccattctac ccactcgaag acgggaccgc cggcgagcag
     4501 ctgcacaaag ccatgaagcg ctacgccctg gtgcccggca ccatcgcctt taccgacgca
     4561 catatcgagg tggacattac ctacgccgag tacttcgaga tgagcgttcg gctggcagaa
     4621 gctatgaagc gctatgggct gaatacaaac catcggatcg tggtgtgcag cgagaatagc
     4681 ttgcagttct tcatgcccgt gttgggtgcc ctgttcatcg gtgtggctgt ggccccagct
     4741 aacgacatct acaacgagcg cgagctgctg aacagcatgg gcatcagcca gcccaccgtc
     4801 gtattcgtga gcaagaaagg gctgcaaaag atcctcaacg tgcaaaagaa gctaccgatc
     4861 atacaaaaga tcatcatcat ggatagcaag accgactacc agggcttcca aagcatgtac
     4921 accttcgtga cttcccattt gccacccggc ttcaacgagt acgacttcgt gcccgagagc
     4981 ttcgaccggg acaaaaccat cgccctgatc atgaacagta gtggcagtac cggattgccc
     5041 aagggcgtag ccctaccgca ccgcaccgct tgtgtccgat tcagtcatgc ccgcgacccc
     5101 atcttcggca accagatcat ccccgacacc gctatcctca gcgtggtgcc atttcaccac
     5161 ggcttcggca tgttcaccac gctgggctac ttgatctgcg gctttcgggt cgtgctcatg
     5221 taccgcttcg aggaggagct attcttgcgc agcttgcaag actataagat tcaatctgcc
     5281 ctgctggtgc ccacactatt tagcttcttc gctaagagca ctctcatcga caagtacgac
     5341 ctaagcaact tgcacgagat cgccagcggc ggggcgccgc tcagcaagga ggtaggtgag
     5401 gccgtggcca aacgcttcca cctaccaggc atccgccagg gctacggcct gacagaaaca
     5461 accagcgcca ttctgatcac ccccgaaggg gacgacaagc ctggcgcagt aggcaaggtg
     5521 gtgcccttct tcgaggctaa ggtggtggac ttggacaccg gtaagacact gggtgtgaac
     5581 cagcgcggcg agctgtgcgt ccgtggcccc atgatcatga gcggctacgt taacaacccc
     5641 gaggctacaa acgctctcat cgacaaggac ggctggctgc acagcggcga catcgcctac
     5701 tgggacgagg acgagcactt cttcatcgtg gaccggctga agagcctgat caaatacaag
     5761 ggctaccagg tagccccagc cgaactggag agcatcctgc tgcaacaccc caacatcttc
     5821 gacgccgggg tcgccggcct gcccgacgac gatgccggcg agctgcccgc cgcagtcgtc
     5881 gtgctggaac acggtaaaac catgaccgag aaggagatcg tggactatgt ggccagccag
     5941 gttacaaccg ccaagaagct gcgcggtggt gttgtgttcg tggacgaggt gcctaaagga
     6001 ctgaccggca agttggacgc ccgcaaaatc cgcgagattc tcattaaggc caagaagggc
     6061 ggcaagatcg ccgtgaaaaa gaccaaagat gaactctaat aattctagag tcggggcggc
     6121 cggccgcttc gagcagacat gataagatac attgatgagt ttggacaaac cacaactaga
     6181 atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc
     6241 attataagct gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt
     6301 cagggggagg tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg tggtaaaatc
     6361 gataaggatc cgtttgcgta ttgggcgctc ttccgctgat ctgcgcagca ccatggcctg
     6421 aaataacctc tgaaagagga acttggttag ctaccttctg aggcggaaag aaccagctgt
     6481 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     6541 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     6601 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     6661 cgcccatccc gcccctaact ccgccc
//
