LOCUS       sequence-182861-        9226 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For dual luciferase report assay.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9226)
  AUTHORS   Zhou W, Pal AS, Hsu AY, Gurol T, Zhu X, Wirbisky-Hershberger SE, 
            Freeman JL, Kasinski AL, Deng Q
  TITLE     MicroRNA-223 Suppresses the Canonical NF-kappaB Pathway in Basal 
            Keratinocytes to Dampen Neutrophilic Inflammation.
  JOURNAL   Cell Rep. 2018 Feb 13;22(7):1810-1823. doi: 
            10.1016/j.celrep.2018.01.058.
  PUBMED    29444433
REFERENCE   2  (bases 1 to 9226)
  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"; doi: 
            "10.1016/j.celrep.2018.01"; journalName: "Cell Rep"; date: 
            "2018-02-13- 13"; volume: "22"; issue: "7"; pages: "1810-1823"
COMMENT     Sequence Label: pSI-check2-Traf6-3'UTR
FEATURES             Location/Qualifiers
     source          1..9226
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      4..592
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     493..512
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        721..1078
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        721..912
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(744..764)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      929..1064
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          1148..1280
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1263..1282
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1325..1343
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1347..1356
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1353..2288
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     polyA_signal    5300..5348
                     /note="synthetic polyadenylation signal"
     promoter        5357..6108
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     regulatory      6138..6147
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             6144..7796
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /db_xref="GI:42718124"
                     /protein_id="AAS38485.1"
                     /translation="MADAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     RBS             7103..7111
                     /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    7840..7961
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7851..7870)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     7905..7924
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        8094..8198
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(8199..8267,8268..9059)
                     /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     complement(8417..8436)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
       61 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
      121 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
      181 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
      241 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
      301 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
      361 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
      421 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
      481 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
      541 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
      601 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tggctcgaca
      661 gatctgcgca gcaccatggc ctgaaataac ctctgaaaga ggaacttggt taggtacctt
      721 ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
      781 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
      841 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
      901 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
      961 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc
     1021 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     1081 gcttgattct tctgacacaa cagtctcgaa cttaagctgc agaagttggt cgtgaggcac
     1141 tgggcaggta agtatcaagg ttacaagaca ggtttaagga gaccaataga aactgggctt
     1201 gtcgagacag agaagactct tgcgtttctg ataggcacct attggtctta ctgacatcca
     1261 ctttgccttt ctctccacag gtgtccactc ccagttcaat tacagctctt aaggctagag
     1321 tacttaatac gactcactat aggctagcca ccatggcttc caaggtgtac gaccccgagc
     1381 aacgcaaacg catgatcact gggcctcagt ggtgggctcg ctgcaagcaa atgaacgtgc
     1441 tggactcctt catcaactac tatgattccg agaagcacgc cgagaacgcc gtgatttttc
     1501 tgcatggtaa cgctgcctcc agctacctgt ggaggcacgt cgtgcctcac atcgagcccg
     1561 tggctagatg catcatccct gatctgatcg gaatgggtaa gtccggcaag agcgggaatg
     1621 gctcatatcg cctcctggat cactacaagt acctcaccgc ttggttcgag ctgctgaacc
     1681 ttccaaagaa aatcatcttt gtgggccacg actggggggc ttgtctggcc tttcactact
     1741 cctacgagca ccaagacaag atcaaggcca tcgtccatgc tgagagtgtc gtggacgtga
     1801 tcgagtcctg ggacgagtgg cctgacatcg aggaggatat cgccctgatc aagagcgaag
     1861 agggcgagaa aatggtgctt gagaataact tcttcgtcga gaccatgctc ccaagcaaga
     1921 tcatgcggaa actggagcct gaggagttcg ctgcctacct ggagccattc aaggagaagg
     1981 gcgaggttag acggcctacc ctctcctggc ctcgcgagat ccctctcgtt aagggaggca
     2041 agcccgacgt cgtccagatt gtccgcaact acaacgccta ccttcgggcc agcgacgatc
     2101 tgcctaagat gttcatcgag tccgaccctg ggttcttttc caacgctatt gtcgagggag
     2161 ctaagaagtt ccctaacacc gagttcgtga aggtgaaggg cctccacttc agccaggagg
     2221 acgctccaga tgaaatgggt aagtacatca agagcttcgt ggagcgcgtg ctgaagaacg
     2281 agcagtaatt ctaggcgatc gctcgaggca ttttttgttt tcagtcttaa gttctcaaag
     2341 acactctgct gaaatgacag aaatacagct aaacctgttt tttttttttt tttgacccat
     2401 gcccatttgt tctgttggtt gaatatcttg ttttatttac aaatgctttc caatgcatat
     2461 ttatgcagtg caatcagtgg acaaatatct gtaggaatca catccccaac tcacaaaata
     2521 ttgtgtcaac caagctaagg gtgtaaaaac tgttgtttta atggtgctgc ttatgctaag
     2581 tgttagcttt cacaaattta attcacgata ctacgtttaa cagctcatac tgccttagaa
     2641 gttgtgtaaa atatatactt ttaaatgaag agatgtctga cagagtacag agcagcactg
     2701 tttattgtga ttgtaggcaa gtttgagcct caaggtcata ggtctcaaac tggattactg
     2761 gagggctgca gctctacaca gttttgctcc acctctaatc aaacactcct catccaacta
     2821 atcaagatgg ttaagactac tcttgaacac cttggctgtt tctcaattcc aagaatgcag
     2881 agaacggact tgtgttcttg tggtgactat tcttccgagg gaattcggca agaacggtca
     2941 ccacaagaac acaagggaga ctgtgagaat agaaaacgcg tcctgtcaga aatgagatgc
     3001 tgtgttcttc actggttctt cctgatggtc acatgacctt cacatgtttt taatgaaaaa
     3061 ttatttaaag attacagcat tcgtacaatg atttattgtt tccccccctt ttcttaatat
     3121 atactatgca taaaatcctt acaagtatac tttgcacaat acaaatgaaa ctcattatgg
     3181 tggcttgagg aagctaacat actgcaatac tacataaact tattattctg cctctgtcgt
     3241 ctcctgagac tagtttctat gtgcatctcc tcctagaccg ttcattctac aaaaaccaaa
     3301 ctaactccga acctcctaac tatactgaat taggttgcta tatcttttcc aactgatcca
     3361 acttacggtt ttccccaaac tgaccctgaa aagtcggaaa agtcccattg acttaacatt
     3421 gtatcaaact ttatgacctc ataactttgt gccagacagt catacagtca ggtttggact
     3481 catttaactc tgactagcaa tctgccaatc actgatgacc tttcaactta ctagccacac
     3541 cctaactacc aattatgaca ccatagcaac tgtcacatag acttccgttg taaagaaaaa
     3601 aaaaaaaaac gtcattgact ttacattgta catactaaca acataccaat tcatgacaaa
     3661 caatatacta gtccatacta gcaacatgct aattcatact ataaacacac tagctacatg
     3721 ctaatttata ctaaaattat gctaaaacat gctagtttat actagaaaca ttctagcaac
     3781 atgccaattc ctacttgaac catgctaatt tatattagta cccacctaga acaccctagc
     3841 aacgccttag cagccacata gcaatcactc agaacacctt agcaaccgcc tagtaaccac
     3901 tcagagcaac cacctagcca gaaacaagct gatccatact agaaacatgc tagcatacat
     3961 ctagttatct ttctatgcca accgtttcaa actttttaaa aactacgtca accttttaga
     4021 caaggctttc tcaagacacc agaaagtttg tccacaaacc tttctagttt taatatgaat
     4081 ttcagcaaat aaacaaccaa cttaatgagt ttatgcctct gttgggattt tcatggtaat
     4141 gtttacttcc actgtcttgt ttagggaaag atacgtttta tctctgacct ttattaataa
     4201 atctatataa aatgctgcgc ttcccacctc atttgtttag ctgcaatgac ttctgggact
     4261 tcacaagcaa gttttgcgtt taagtctgca atgtcgtgtc cacgatatca agaacacatc
     4321 cgggaaattt catgtgtcct ccattcttgc ggtcttgagt ttaggaactg aactttggca
     4381 gttgatgatg atgttacagg aggacacaag atcactaaag aacccatatt gagaaacagc
     4441 ctttggttag ttggatcaga tgtgtttgaa tagggttgga gcaaaactgc agagctgtgg
     4501 ccctccagga attgagtttg agacctatgc tctaggtcat taacagcttt aaaatgttta
     4561 tagacataat ggggctatat gtagaattca gaattcattt tgttgttgaa gttgaaaata
     4621 tttaccgtct atgcttcaaa ccaatatact gataatgaac atcaacgtca ttcgtgaggc
     4681 taagagtgtg tgctttcttc tcaacaaaat aaacacagca aaatttacca cagtgagaaa
     4741 actgctgtga agaaagcatc tgaccatagc agtgtcgatt ggttgaaagt tctgtaattc
     4801 accagcgtct tgtcgacaga tgaggtaata aatattgaat tgacacatga aagttaagct
     4861 aataaagcct tgttcacttc aatatgttta atttataatg cctcaaagga caacctccaa
     4921 atggagaatt ttttttatat tgtccataag gtatttacac aatggcagta aaagcattta
     4981 gtgcttaaac attattacat ttaccccttt aaggtgtgat catttcagca acatttcact
     5041 gaaaattcat tttcactagg tttttaatct atggtcccaa cagctttctg tcatctaatg
     5101 ctgctcattt gaagtcattc tgatatcaga ctgggtaaat ctttcaccct cacatcaaat
     5161 ttccagttgg gtatattgtg atgactggat ttcagagaca aaacctagca caacaacatt
     5221 tctgaaaagg gttgtatgta ctaatctcat gtcctataaa taataatgtt ttaatgtgcg
     5281 gccgctggcc gctggccgca ataaaatatc tttattttca ttacatctgt gtgttggttt
     5341 tttgtgtgag gatctaaatg agtcttcgga cctcgcgggg gccgcttaag cggtggttag
     5401 ggtttgtctg acgcgggggg agggggaagg aacgaaacac tctcattcgg aggcggctcg
     5461 gggtttggtc ttggtggcca cgggcacgca gaagagcgcc gcgatcctct taagcacccc
     5521 cccgccctcc gtggaggcgg gggtttggtc ggcgggtggt aactggcggg ccgctgactc
     5581 gggcgggtcg cgcgccccag agtgtgacct tttcggtctg ctcgcagacc cccgggcggc
     5641 gccgccgcgg cggcgacggg ctcgctgggt cctaggctcc atggggaccg tatacgtgga
     5701 caggctctgg agcatccgca cgactgcggt gatattaccg gagaccttct gcgggacgag
     5761 ccgggtcacg cggctgacgc ggagcgtccg ttgggcgaca aacaccagga cggggcacag
     5821 gtacactatc ttgtcacccg gaggcgcgag ggactgcagg agcttcaggg agtggcgcag
     5881 ctgcttcatc cccgtggccc gttgctcgcg tttgctggcg gtgtccccgg aagaaatata
     5941 tttgcatgtc tttagttcta tgatgacaca aaccccgccc agcgtcttgt cattggcgaa
     6001 ttcgaacacg cagatgcagt cggggcggcg cggtcccagg tccacttcgc atattaaggt
     6061 gacgcgtgtg gcctcgaaca ccgagcgacc ctgcagcgac ccgcttaaaa gcttggcatt
     6121 ccggtactgt tggtaaagcc accatggccg atgctaagaa cattaagaag ggccctgctc
     6181 ccttctaccc tctggaggat ggcaccgctg gcgagcagct gcacaaggcc atgaagaggt
     6241 atgccctggt gcctggcacc attgccttca ccgatgccca cattgaggtg gacatcacct
     6301 atgccgagta cttcgagatg tctgtgcgcc tggccgaggc catgaagagg tacggcctga
     6361 acaccaacca ccgcatcgtg gtgtgctctg agaactctct gcagttcttc atgccagtgc
     6421 tgggcgccct gttcatcgga gtggccgtgg cccctgctaa cgacatttac aacgagcgcg
     6481 agctgctgaa cagcatgggc atttctcagc ctaccgtggt gttcgtgtct aagaagggcc
     6541 tgcagaagat cctgaacgtg cagaagaagc tgcctatcat ccagaagatc atcatcatgg
     6601 actctaagac cgactaccag ggcttccaga gcatgtacac attcgtgaca tctcatctgc
     6661 ctcctggctt caacgagtac gacttcgtgc cagagtcttt cgacagggac aaaaccattg
     6721 ccctgatcat gaacagctct gggtctaccg gcctgcctaa gggcgtggcc ctgcctcatc
     6781 gcaccgcctg tgtgcgcttc tctcacgccc gcgaccctat tttcggcaac cagatcatcc
     6841 ccgacaccgc tattctgagc gtggtgccat tccaccacgg cttcggcatg ttcaccaccc
     6901 tgggctacct gatttgcggc tttcgggtgg tgctgatgta ccgcttcgag gaggagctgt
     6961 tcctgcgcag cctgcaagac tacaaaattc agtctgccct gctggtgcca accctgttca
     7021 gcttcttcgc taagagcacc ctgatcgaca agtacgacct gtctaacctg cacgagattg
     7081 cctctggcgg cgccccactg tctaaggagg tgggcgaagc cgtggccaag cgctttcatc
     7141 tgccaggcat ccgccagggc tacggcctga ccgagacaac cagcgccatt ctgattaccc
     7201 cagagggcga cgacaagcct ggcgccgtgg gcaaggtggt gccattcttc gaggccaagg
     7261 tggtggacct ggacaccggc aagaccctgg gagtgaacca gcgcggcgag ctgtgtgtgc
     7321 gcggccctat gattatgtcc ggctacgtga ataaccctga ggccacaaac gccctgatcg
     7381 acaaggacgg ctggctgcac tctggcgaca ttgcctactg ggacgaggac gagcacttct
     7441 tcatcgtgga ccgcctgaag tctctgatca agtacaaggg ctaccaggtg gccccagccg
     7501 agctggagtc tatcctgctg cagcacccta acattttcga cgccggagtg gccggcctgc
     7561 ccgacgacga tgccggcgag ctgcctgccg ccgtcgtcgt gctggaacac ggcaagacca
     7621 tgaccgagaa ggagatcgtg gactatgtgg ccagccaggt gacaaccgcc aagaagctgc
     7681 gcggcggagt ggtgttcgtg gacgaggtgc ccaagggcct gaccggcaag ctggacgccc
     7741 gcaagatccg cgagatcctg atcaaggcta agaaaggcgg caagatcgcc gtgtaataat
     7801 tctagagtcg gggcggccgg ccgcttcgag cagacatgat aagatacatt gatgagtttg
     7861 gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta
     7921 ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc
     7981 attttatgtt tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag taaaacctct
     8041 acaaatgtgg taaaatcgat aaggatccag gtggcacttt tcggggaaat gtgcgcggaa
     8101 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     8161 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     8221 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     8281 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     8341 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     8401 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     8461 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     8521 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     8581 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     8641 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     8701 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     8761 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     8821 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     8881 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     8941 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     9001 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     9061 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     9121 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     9181 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatc
//
