LOCUS       sequence-352603-        6836 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 with an HA tag and the 
            eleventh beta-strand of GFP added to the C-terminus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6836)
  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 6836)
  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-HA-GFP11-KDEL
FEATURES             Location/Qualifiers
     source          1..6836
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             81..1118
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             1097..1105
                     /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    1142..1190
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(1346..1363)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1517..2105)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1597..1616)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2305..3096,3097..3165))
                     /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    3270..3318
                     /note="synthetic polyadenylation signal"
     misc_feature    3332..3423
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     3372..3391
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     enhancer        3574..3953
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3954..4157
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4108..4128
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4154..4178
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             4350..5996
                     /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             6033..6059
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             6099..6146
                     /codon_start=1
                     /product="11th beta-strand of superfolder GFP"
                     /label=GFP11
                     /note="associates with GFP(1-10), consisting of the first 
                     10 beta-strands, to reconstitute fluorescent GFP (Cabantous
                     et al., 2005; Kamiyama et al., 2016)"
                     /translation="RDHMVLHEYVNAAGIT"
     polyA_signal    6214..6335
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6225..6244)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6279..6298
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        join(6529..6836,1..50)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        6529..6720
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(6552..6572)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      join(6737..6836,1..36)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6799..6818
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
ORIGIN
        1 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcgatt
       61 cttctgacac tagcgccacc atgaagaagc ccgaactcac cgctaccagc gttgaaaaat
      121 ttctcatcga gaagttcgac agtgtgagcg acctgatgca gttgtcggag ggcgaagaga
      181 gccgagcctt cagcttcgat gtcggcggac gcggctatgt actgcgggtg aatagctgcg
      241 ctgatggctt ctacaaagac cgctacgtgt accgccactt cgccagcgct gcactaccca
      301 tccccgaagt gttggacatc ggcgagttca gcgagagcct gacatactgc atcagtagac
      361 gcgcccaagg cgttactctc caagacctcc ccgaaacaga gctgcctgct gtgttacagc
      421 ctgtcgccga agctatggat gctattgccg ccgccgacct cagtcaaacc agcggcttcg
      481 gcccattcgg gccccaaggc atcggccagt acacaacctg gcgggatttc atttgcgcca
      541 ttgctgatcc ccatgtctac cactggcaga ccgtgatgga cgacaccgtg tccgccagcg
      601 tagctcaagc cctggacgaa ctgatgctgt gggccgaaga ctgtcccgag gtgcgccacc
      661 tcgtccatgc cgacttcggc agcaacaacg tcctgaccga caacggccgc atcaccgccg
      721 taatcgactg gtccgaagct atgttcgggg acagtcagta cgaggtggcc aacatcttct
      781 tctggcggcc ctggctggct tgcatggagc agcagactcg ctacttcgag cgccggcatc
      841 ccgagctggc cggcagccct cgtctgcgag cctacatgct gcgcatcggc ctggatcagc
      901 tctaccagag cctcgtggac ggcaacttcg acgatgctgc ctgggctcaa ggccgctgcg
      961 atgccatcgt ccgcagcggg gccggcaccg tcggtcgcac acaaatcgct cgccggagcg
     1021 cagccgtatg gaccgacggc tgcgtcgagg tgctggccga cagcggcaac cgccggccca
     1081 gtacacgacc gcgcgctaag gaggtaggtc gagtttaaac tctagaaccg gtcatggccg
     1141 caataaaata tctttatttt cattacatct gtgtgttggt tttttgtgtg ttcgaactag
     1201 atgctgtcga ccgatgccct tgagagcctt caacccagtc agctccttcc ggtgggcgcg
     1261 gggcatgact atcgtcgccg cacttatgac tgtcttcttt atcatgcaac tcgtaggaca
     1321 ggtgccggca gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     1381 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     1441 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     1501 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     1561 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     1621 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     1681 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     1741 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     1801 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     1861 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     1921 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     1981 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     2041 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     2101 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     2161 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     2221 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagcggcc
     2281 gcaaatgcta aaccactgca gtggttacca gtgcttgatc agtgaggcac cgatctcagc
     2341 gatctgccta tttcgttcgt ccatagtggc ctgactcccc gtcgtgtaga tcactacgat
     2401 tcgtgagggc ttaccatcag gccccagcgc agcaatgatg ccgcgagagc cgcgttcacc
     2461 ggcccccgat ttgtcagcaa tgaaccagcc agcagggagg gccgagcgaa gaagtggtcc
     2521 tgctactttg tccgcctcca tccagtctat gagctgctgt cgtgatgcta gagtaagaag
     2581 ttcgccagtg agtagtttcc gaagagttgt ggccattgct actggcatcg tggtatcacg
     2641 ctcgtcgttc ggtatggctt cgttcaactc tggttcccag cggtcaagcc gggtcacatg
     2701 atcacccata ttatgaagaa atgcagtcag ctccttaggg cctccgatcg ttgtcagaag
     2761 taagttggcc gcggtgttgt cgctcatggt aatggcagca ctacacaatt ctcttaccgt
     2821 catgccatcc gtaagatgct tttccgtgac cggcgagtac tcaaccaagt cgttttgtga
     2881 gtagtgtata cggcgaccaa gctgctcttg cccggcgtct atacgggaca acaccgcgcc
     2941 acatagcagt actttgaaag tgctcatcat cgggaatcgt tcttcggggc ggaaagactc
     3001 aaggatcttg ccgctattga gatccagttc gatatagccc actcttgcac ccagttgatc
     3061 ttcagcatct tttactttca ccagcgtttc ggggtgtgca aaaacaggca agcaaaatgc
     3121 cgcaaagaag ggaatgagtg cgacacgaaa atgttggatg ctcatactcg tcctttttca
     3181 atattattga agcatttatc agggttacta gtacgtctct caaggataag taagtaatat
     3241 taaggtacgg gaggtattgg acaggccgca ataaaatatc tttattttca ttacatctgt
     3301 gtgttggttt tttgtgtgaa tcgatagtac taacatacgc tctccatcaa aacaaaacga
     3361 aacaaaacaa actagcaaaa taggctgtcc ccagtgcaag tgcaggtgcc agaacatttc
     3421 tctggcctaa ctggcctcaa tattggccat tagccatatt attcattggt tatatagcat
     3481 aaatcaatat tggctattgg ccattgcata cgttgtatct atatcataat atgtacattt
     3541 atattggctc atgtccaata tgaccgccat gttggcattg attattgact agttattaat
     3601 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac
     3661 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa
     3721 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
     3781 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtccgcccc
     3841 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttac
     3901 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc
     3961 ggttttggca gtacaccaat gggcgtggat agcggtttga ctcacgggga tttccaagtc
     4021 tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa
     4081 aatgtcgtaa taaccccgcc ccgttgacgc aaatgggcgg taggcgtgta cggtgggagg
     4141 tctatataag cagagctcgt ttagtgaacc gtcagatcac tagaagcttg gcaacagtac
     4201 cgtgatacaa ccacactcga gccatgaaca tcaaaggatc gccatggaaa gggtccctcc
     4261 tgctgctgct ggtgtcaaac ctgctcctgt gccagagcgt ggccccctgg aattcagctt
     4321 ggcaatccgg tactgttggt aaagccaccg aagatgccaa aaacattaag aagggcccag
     4381 cgccattcta cccactcgaa gacgggaccg ccggcgagca gctgcacaaa gccatgaagc
     4441 gctacgccct ggtgcccggc accatcgcct ttaccgacgc acatatcgag gtggacatta
     4501 cctacgccga gtacttcgag atgagcgttc ggctggcaga agctatgaag cgctatgggc
     4561 tgaatacaaa ccatcggatc gtggtgtgca gcgagaatag cttgcagttc ttcatgcccg
     4621 tgttgggtgc cctgttcatc ggtgtggctg tggccccagc taacgacatc tacaacgagc
     4681 gcgagctgct gaacagcatg ggcatcagcc agcccaccgt cgtattcgtg agcaagaaag
     4741 ggctgcaaaa gatcctcaac gtgcaaaaga agctaccgat catacaaaag atcatcatca
     4801 tggatagcaa gaccgactac cagggcttcc aaagcatgta caccttcgtg acttcccatt
     4861 tgccacccgg cttcaacgag tacgacttcg tgcccgagag cttcgaccgg gacaaaacca
     4921 tcgccctgat catgaacagt agtggcagta ccggattgcc caagggcgta gccctaccgc
     4981 accgcaccgc ttgtgtccga ttcagtcatg cccgcgaccc catcttcggc aaccagatca
     5041 tccccgacac cgctatcctc agcgtggtgc catttcacca cggcttcggc atgttcacca
     5101 cgctgggcta cttgatctgc ggctttcggg tcgtgctcat gtaccgcttc gaggaggagc
     5161 tattcttgcg cagcttgcaa gactataaga ttcaatctgc cctgctggtg cccacactat
     5221 ttagcttctt cgctaagagc actctcatcg acaagtacga cctaagcaac ttgcacgaga
     5281 tcgccagcgg cggggcgccg ctcagcaagg aggtaggtga ggccgtggcc aaacgcttcc
     5341 acctaccagg catccgccag ggctacggcc tgacagaaac aaccagcgcc attctgatca
     5401 cccccgaagg ggacgacaag cctggcgcag taggcaaggt ggtgcccttc ttcgaggcta
     5461 aggtggtgga cttggacacc ggtaagacac tgggtgtgaa ccagcgcggc gagctgtgcg
     5521 tccgtggccc catgatcatg agcggctacg ttaacaaccc cgaggctaca aacgctctca
     5581 tcgacaagga cggctggctg cacagcggcg acatcgccta ctgggacgag gacgagcact
     5641 tcttcatcgt ggaccggctg aagagcctga tcaaatacaa gggctaccag gtagccccag
     5701 ccgaactgga gagcatcctg ctgcaacacc ccaacatctt cgacgccggg gtcgccggcc
     5761 tgcccgacga cgatgccggc gagctgcccg ccgcagtcgt cgtgctggaa cacggtaaaa
     5821 ccatgaccga gaaggagatc gtggactatg tggccagcca ggttacaacc gccaagaagc
     5881 tgcgcggtgg tgttgtgttc gtggacgagg tgcctaaagg actgaccggc aagttggacg
     5941 cccgcaagat ccgcgagatt ctcattaagg ccaagaaggg cggcaagatc gccgtgatcc
     6001 ccgggggatc gggtgacggt ccggtcgcca cctacccata cgatgttcca gattacgctg
     6061 gtcgacggat cggtggtggt tctggaggcg gttctatgag agatcatatg gttttgcatg
     6121 aatatgttaa tgctgctggt attactaaaa agaccaaaga tgaactctaa taattctaga
     6181 gtcggggcgg ccggccgctt cgagcagaca tgataagata cattgatgag tttggacaaa
     6241 ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt
     6301 tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta
     6361 tgtttcaggt tcagggggag gtgtgggagg ttttttaaag caagtaaaac ctctacaaat
     6421 gtggtaaaat cgataaggat ccgtttgcgt attgggcgct cttccgctga tctgcgcagc
     6481 accatggcct gaaataacct ctgaaagagg aacttggtta gctaccttct gaggcggaaa
     6541 gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg
     6601 cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg
     6661 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc
     6721 gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca
     6781 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct ctgagc
//
