LOCUS       sequence-352618-        6836 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  DM 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-DM-HA-GFP11-KDEL
FEATURES             Location/Qualifiers
     source          1..6836
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             62..1099
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             1078..1086
                     /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    1123..1171
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(1327..1344)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1498..2086)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1578..1597)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2286..3077,3078..3146))
                     /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    3251..3299
                     /note="synthetic polyadenylation signal"
     misc_feature    3313..3404
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     3353..3372
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     enhancer        3555..3934
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3935..4138
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4089..4109
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4135..4159
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             4331..5977
                     /codon_start=1
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="synthetic luc2 version of the luciferase gene"
                     /translation="EDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDAH
                     IEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAPA
                     NDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQSM
                     YTFVTSHLPPGFNEYDFVPESFDQDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHAR
                     DPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFQVVLMYRFEEELFLRSLQDYKI
                     QSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGYG
                     LTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMSG
                     YVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESILL
                     QHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVVF
                     VDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     CDS             6014..6040
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             6080..6127
                     /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    6195..6316
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6206..6225)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6260..6279
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        join(6510..6836,1..31)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        6510..6701
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(6533..6553)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      join(6718..6836,1..17)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6780..6799
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
ORIGIN
        1 aggctttttt ggaggcctag gcttttgcaa aaagctcgat tcttctgaca ctagcgccac
       61 catgaagaag cccgaactca ccgctaccag cgttgaaaaa tttctcatcg agaagttcga
      121 cagtgtgagc gacctgatgc agttgtcgga gggcgaagag agccgagcct tcagcttcga
      181 tgtcggcgga cgcggctatg tactgcgggt gaatagctgc gctgatggct tctacaaaga
      241 ccgctacgtg taccgccact tcgccagcgc tgcactaccc atccccgaag tgttggacat
      301 cggcgagttc agcgagagcc tgacatactg catcagtaga cgcgcccaag gcgttactct
      361 ccaagacctc cccgaaacag agctgcctgc tgtgttacag cctgtcgccg aagctatgga
      421 tgctattgcc gccgccgacc tcagtcaaac cagcggcttc ggcccattcg ggccccaagg
      481 catcggccag tacacaacct ggcgggattt catttgcgcc attgctgatc cccatgtcta
      541 ccactggcag accgtgatgg acgacaccgt gtccgccagc gtagctcaag ccctggacga
      601 actgatgctg tgggccgaag actgtcccga ggtgcgccac ctcgtccatg ccgacttcgg
      661 cagcaacaac gtcctgaccg acaacggccg catcaccgcc gtaatcgact ggtccgaagc
      721 tatgttcggg gacagtcagt acgaggtggc caacatcttc ttctggcggc cctggctggc
      781 ttgcatggag cagcagactc gctacttcga gcgccggcat cccgagctgg ccggcagccc
      841 tcgtctgcga gcctacatgc tgcgcatcgg cctggatcag ctctaccaga gcctcgtgga
      901 cggcaacttc gacgatgctg cctgggctca aggccgctgc gatgccatcg tccgcagcgg
      961 ggccggcacc gtcggtcgca cacaaatcgc tcgccggagc gcagccgtat ggaccgacgg
     1021 ctgcgtcgag gtgctggccg acagcggcaa ccgccggccc agtacacgac cgcgcgctaa
     1081 ggaggtaggt cgagtttaaa ctctagaacc ggtcatggcc gcaataaaat atctttattt
     1141 tcattacatc tgtgtgttgg ttttttgtgt gttcgaacta gatgctgtcg accgatgccc
     1201 ttgagagcct tcaacccagt cagctccttc cggtgggcgc ggggcatgac tatcgtcgcc
     1261 gcacttatga ctgtcttctt tatcatgcaa ctcgtaggac aggtgccggc agcgctcttc
     1321 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     1381 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     1441 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     1501 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     1561 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     1621 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     1681 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     1741 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     1801 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     1861 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     1921 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     1981 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     2041 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     2101 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     2161 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     2221 aatctaaagt atatatgagt aaacttggtc tgacagcggc cgcaaatgct aaaccactgc
     2281 agtggttacc agtgcttgat cagtgaggca ccgatctcag cgatctgcct atttcgttcg
     2341 tccatagtgg cctgactccc cgtcgtgtag atcactacga ttcgtgaggg cttaccatca
     2401 ggccccagcg cagcaatgat gccgcgagag ccgcgttcac cggcccccga tttgtcagca
     2461 atgaaccagc cagcagggag ggccgagcga agaagtggtc ctgctacttt gtccgcctcc
     2521 atccagtcta tgagctgctg tcgtgatgct agagtaagaa gttcgccagt gagtagtttc
     2581 cgaagagttg tggccattgc tactggcatc gtggtatcac gctcgtcgtt cggtatggct
     2641 tcgttcaact ctggttccca gcggtcaagc cgggtcacat gatcacccat attatgaaga
     2701 aatgcagtca gctccttagg gcctccgatc gttgtcagaa gtaagttggc cgcggtgttg
     2761 tcgctcatgg taatggcagc actacacaat tctcttaccg tcatgccatc cgtaagatgc
     2821 ttttccgtga ccggcgagta ctcaaccaag tcgttttgtg agtagtgtat acggcgacca
     2881 agctgctctt gcccggcgtc tatacgggac aacaccgcgc cacatagcag tactttgaaa
     2941 gtgctcatca tcgggaatcg ttcttcgggg cggaaagact caaggatctt gccgctattg
     3001 agatccagtt cgatatagcc cactcttgca cccagttgat cttcagcatc ttttactttc
     3061 accagcgttt cggggtgtgc aaaaacaggc aagcaaaatg ccgcaaagaa gggaatgagt
     3121 gcgacacgaa aatgttggat gctcatactc gtcctttttc aatattattg aagcatttat
     3181 cagggttact agtacgtctc tcaaggataa gtaagtaata ttaaggtacg ggaggtattg
     3241 gacaggccgc aataaaatat ctttattttc attacatctg tgtgttggtt ttttgtgtga
     3301 atcgatagta ctaacatacg ctctccatca aaacaaaacg aaacaaaaca aactagcaaa
     3361 ataggctgtc cccagtgcaa gtgcaggtgc cagaacattt ctctggccta actggcctca
     3421 atattggcca ttagccatat tattcattgg ttatatagca taaatcaata ttggctattg
     3481 gccattgcat acgttgtatc tatatcataa tatgtacatt tatattggct catgtccaat
     3541 atgaccgcca tgttggcatt gattattgac tagttattaa tagtaatcaa ttacggggtc
     3601 attagttcat agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc
     3661 tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt
     3721 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca
     3781 cttggcagta catcaagtgt atcatatgcc aagtccgccc cctattgacg tcaatgacgg
     3841 taaatggccc gcctggcatt atgcccagta catgacctta cgggactttc ctacttggca
     3901 gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc agtacaccaa
     3961 tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa
     4021 tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta ataaccccgc
     4081 cccgttgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctcg
     4141 tttagtgaac cgtcagatca ctagaagctt ggcaacagta ccgtgataca accacactcg
     4201 agccatgaac atcaaaggat cgccatggaa agggtccctc ctgctgctgc tggtgtcaaa
     4261 cctgctcctg tgccagagcg tggccccctg gaattcagct tggcaatccg gtactgttgg
     4321 taaagccacc gaagatgcca aaaacattaa gaagggccca gcgccattct acccactcga
     4381 agacgggacc gccggcgagc agctgcacaa agccatgaag cgctacgccc tggtgcccgg
     4441 caccatcgcc tttaccgacg cacatatcga ggtggacatt acctacgccg agtacttcga
     4501 gatgagcgtt cggctggcag aagctatgaa gcgctatggg ctgaatacaa accatcggat
     4561 cgtggtgtgc agcgagaata gcttgcagtt cttcatgccc gtgttgggtg ccctgttcat
     4621 cggtgtggct gtggccccag ctaacgacat ctacaacgag cgcgagctgc tgaacagcat
     4681 gggcatcagc cagcccaccg tcgtattcgt gagcaagaaa gggctgcaaa agatcctcaa
     4741 cgtgcaaaag aagctaccga tcatacaaaa gatcatcatc atggatagca agaccgacta
     4801 ccagggcttc caaagcatgt acaccttcgt gacttcccat ttgccacccg gcttcaacga
     4861 gtacgacttc gtgcccgaga gcttcgacca ggacaaaacc atcgccctga tcatgaacag
     4921 tagtggcagt accggattgc ccaagggcgt agccctaccg caccgcaccg cttgtgtccg
     4981 attcagtcat gcccgcgacc ccatcttcgg caaccagatc atccccgaca ccgctatcct
     5041 cagcgtggtg ccatttcacc acggcttcgg catgttcacc acgctgggct acttgatctg
     5101 cggctttcaa gtcgtgctca tgtaccgctt cgaggaggag ctattcttgc gcagcttgca
     5161 agactataag attcaatctg ccctgctggt gcccacacta tttagcttct tcgctaagag
     5221 cactctcatc gacaagtacg acctaagcaa cttgcacgag atcgccagcg gcggggcgcc
     5281 gctcagcaag gaggtaggtg aggccgtggc caaacgcttc cacctaccag gcatccgcca
     5341 gggctacggc ctgacagaaa caaccagcgc cattctgatc acccccgaag gggacgacaa
     5401 gcctggcgca gtaggcaagg tggtgccctt cttcgaggct aaggtggtgg acttggacac
     5461 cggtaagaca ctgggtgtga accagcgcgg cgagctgtgc gtccgtggcc ccatgatcat
     5521 gagcggctac gttaacaacc ccgaggctac aaacgctctc atcgacaagg acggctggct
     5581 gcacagcggc gacatcgcct actgggacga ggacgagcac ttcttcatcg tggaccggct
     5641 gaagagcctg atcaaataca agggctacca ggtagcccca gccgaactgg agagcatcct
     5701 gctgcaacac cccaacatct tcgacgccgg ggtcgccggc ctgcccgacg acgatgccgg
     5761 cgagctgccc gccgcagtcg tcgtgctgga acacggtaaa accatgaccg agaaggagat
     5821 cgtggactat gtggccagcc aggttacaac cgccaagaag ctgcgcggtg gtgttgtgtt
     5881 cgtggacgag gtgcctaaag gactgaccgg caagttggac gcccgcaaga tccgcgagat
     5941 tctcattaag gccaagaagg gcggcaagat cgccgtgatc cccgggggat cgggtgacgg
     6001 tccggtcgcc acctacccat acgatgttcc agattacgct ggtcgacgga tcggtggtgg
     6061 ttctggaggc ggttctatga gagatcatat ggttttgcat gaatatgtta atgctgctgg
     6121 tattactaaa aagaccaaag atgaactcta ataattctag agtcggggcg gccggccgct
     6181 tcgagcagac atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg
     6241 aaaaaaatgc tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag
     6301 ctgcaataaa caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga
     6361 ggtgtgggag gttttttaaa gcaagtaaaa cctctacaaa tgtggtaaaa tcgataagga
     6421 tccgtttgcg tattgggcgc tcttccgctg atctgcgcag caccatggcc tgaaataacc
     6481 tctgaaagag gaacttggtt agctaccttc tgaggcggaa agaaccagct gtggaatgtg
     6541 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     6601 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
     6661 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     6721 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     6781 atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgagg
//
