LOCUS       sequence-322208-        6271 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression vector, Luciferase reporter to test activity of
            miRNA (miR-15-5p).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6271)
  TITLE     Karreth lab plasmids
REFERENCE   2  (bases 1 to 6271)
  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"
COMMENT     Sequence Label: psiCHECK2-miR-15-5p
FEATURES             Location/Qualifiers
     source          1..6271
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        66..423
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        66..257
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(89..109)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      274..409
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          493..625
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     608..627
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        670..688
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      692..701
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             698..1633
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     polyA_signal    1690..1738
                     /note="synthetic polyadenylation signal"
     promoter        1747..2498
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     regulatory      2528..2537
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2534..4186
                     /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             3493..3501
                     /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    4230..4351
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4241..4260)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4295..4314
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        4484..4588
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4589..4657,4658..5449)
                     /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(4807..4826)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5620..6208
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6109..6128
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgacagatct gcgcagcacc atggcctgaa ataacctctg aaagaggaac ttggttaggt
       61 accttctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc
      121 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
      181 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
      241 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
      301 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
      361 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
      421 aaaaagcttg attcttctga cacaacagtc tcgaacttaa gctgcagaag ttggtcgtga
      481 ggcactgggc aggtaagtat caaggttaca agacaggttt aaggagacca atagaaactg
      541 ggcttgtcga gacagagaag actcttgcgt ttctgatagg cacctattgg tcttactgac
      601 atccactttg cctttctctc cacaggtgtc cactcccagt tcaattacag ctcttaaggc
      661 tagagtactt aatacgactc actataggct agccaccatg gcttccaagg tgtacgaccc
      721 cgagcaacgc aaacgcatga tcactgggcc tcagtggtgg gctcgctgca agcaaatgaa
      781 cgtgctggac tccttcatca actactatga ttccgagaag cacgccgaga acgccgtgat
      841 ttttctgcat ggtaacgctg cctccagcta cctgtggagg cacgtcgtgc ctcacatcga
      901 gcccgtggct agatgcatca tccctgatct gatcggaatg ggtaagtccg gcaagagcgg
      961 gaatggctca tatcgcctcc tggatcacta caagtacctc accgcttggt tcgagctgct
     1021 gaaccttcca aagaaaatca tctttgtggg ccacgactgg ggggcttgtc tggcctttca
     1081 ctactcctac gagcaccaag acaagatcaa ggccatcgtc catgctgaga gtgtcgtgga
     1141 cgtgatcgag tcctgggacg agtggcctga catcgaggag gatatcgccc tgatcaagag
     1201 cgaagagggc gagaaaatgg tgcttgagaa taacttcttc gtcgagacca tgctcccaag
     1261 caagatcatg cggaaactgg agcctgagga gttcgctgcc tacctggagc cattcaagga
     1321 gaagggcgag gttagacggc ctaccctctc ctggcctcgc gagatccctc tcgttaaggg
     1381 aggcaagccc gacgtcgtcc agattgtccg caactacaac gcctaccttc gggccagcga
     1441 cgatctgcct aagatgttca tcgagtccga ccctgggttc ttttccaacg ctattgtcga
     1501 gggagctaag aagttcccta acaccgagtt cgtgaaggtg aagggcctcc acttcagcca
     1561 ggaggacgct ccagatgaaa tgggtaagta catcaagagc ttcgtggagc gcgtgctgaa
     1621 gaacgagcag taattctagg cgatcgctcg agcacaaacc attatgtgct gctagcggcc
     1681 gctggccgca ataaaatatc tttattttca ttacatctgt gtgttggttt tttgtgtgag
     1741 gatctaaatg agtcttcgga cctcgcgggg gccgcttaag cggtggttag ggtttgtctg
     1801 acgcgggggg agggggaagg aacgaaacac tctcattcgg aggcggctcg gggtttggtc
     1861 ttggtggcca cgggcacgca gaagagcgcc gcgatcctct taagcacccc cccgccctcc
     1921 gtggaggcgg gggtttggtc ggcgggtggt aactggcggg ccgctgactc gggcgggtcg
     1981 cgcgccccag agtgtgacct tttcggtctg ctcgcagacc cccgggcggc gccgccgcgg
     2041 cggcgacggg ctcgctgggt cctaggctcc atggggaccg tatacgtgga caggctctgg
     2101 agcatccgca cgactgcggt gatattaccg gagaccttct gcgggacgag ccgggtcacg
     2161 cggctgacgc ggagcgtccg ttgggcgaca aacaccagga cggggcacag gtacactatc
     2221 ttgtcacccg gaggcgcgag ggactgcagg agcttcaggg agtggcgcag ctgcttcatc
     2281 cccgtggccc gttgctcgcg tttgctggcg gtgtccccgg aagaaatata tttgcatgtc
     2341 tttagttcta tgatgacaca aaccccgccc agcgtcttgt cattggcgaa ttcgaacacg
     2401 cagatgcagt cggggcggcg cggtcccagg tccacttcgc atattaaggt gacgcgtgtg
     2461 gcctcgaaca ccgagcgacc ctgcagcgac ccgcttaaaa gcttggcatt ccggtactgt
     2521 tggtaaagcc accatggccg atgctaagaa cattaagaag ggccctgctc ccttctaccc
     2581 tctggaggat ggcaccgctg gcgagcagct gcacaaggcc atgaagaggt atgccctggt
     2641 gcctggcacc attgccttca ccgatgccca cattgaggtg gacatcacct atgccgagta
     2701 cttcgagatg tctgtgcgcc tggccgaggc catgaagagg tacggcctga acaccaacca
     2761 ccgcatcgtg gtgtgctctg agaactctct gcagttcttc atgccagtgc tgggcgccct
     2821 gttcatcgga gtggccgtgg cccctgctaa cgacatttac aacgagcgcg agctgctgaa
     2881 cagcatgggc atttctcagc ctaccgtggt gttcgtgtct aagaagggcc tgcagaagat
     2941 cctgaacgtg cagaagaagc tgcctatcat ccagaagatc atcatcatgg actctaagac
     3001 cgactaccag ggcttccaga gcatgtacac attcgtgaca tctcatctgc ctcctggctt
     3061 caacgagtac gacttcgtgc cagagtcttt cgacagggac aaaaccattg ccctgatcat
     3121 gaacagctct gggtctaccg gcctgcctaa gggcgtggcc ctgcctcatc gcaccgcctg
     3181 tgtgcgcttc tctcacgccc gcgaccctat tttcggcaac cagatcatcc ccgacaccgc
     3241 tattctgagc gtggtgccat tccaccacgg cttcggcatg ttcaccaccc tgggctacct
     3301 gatttgcggc tttcgggtgg tgctgatgta ccgcttcgag gaggagctgt tcctgcgcag
     3361 cctgcaagac tacaaaattc agtctgccct gctggtgcca accctgttca gcttcttcgc
     3421 taagagcacc ctgatcgaca agtacgacct gtctaacctg cacgagattg cctctggcgg
     3481 cgccccactg tctaaggagg tgggcgaagc cgtggccaag cgctttcatc tgccaggcat
     3541 ccgccagggc tacggcctga ccgagacaac cagcgccatt ctgattaccc cagagggcga
     3601 cgacaagcct ggcgccgtgg gcaaggtggt gccattcttc gaggccaagg tggtggacct
     3661 ggacaccggc aagaccctgg gagtgaacca gcgcggcgag ctgtgtgtgc gcggccctat
     3721 gattatgtcc ggctacgtga ataaccctga ggccacaaac gccctgatcg acaaggacgg
     3781 ctggctgcac tctggcgaca ttgcctactg ggacgaggac gagcacttct tcatcgtgga
     3841 ccgcctgaag tctctgatca agtacaaggg ctaccaggtg gccccagccg agctggagtc
     3901 tatcctgctg cagcacccta acattttcga cgccggagtg gccggcctgc ccgacgacga
     3961 tgccggcgag ctgcctgccg ccgtcgtcgt gctggaacac ggcaagacca tgaccgagaa
     4021 ggagatcgtg gactatgtgg ccagccaggt gacaaccgcc aagaagctgc gcggcggagt
     4081 ggtgttcgtg gacgaggtgc ccaagggcct gaccggcaag ctggacgccc gcaagatccg
     4141 cgagatcctg atcaaggcta agaaaggcgg caagatcgcc gtgtaataat tctagagtcg
     4201 gggcggccgg ccgcttcgag cagacatgat aagatacatt gatgagtttg gacaaaccac
     4261 aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt
     4321 tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc attttatgtt
     4381 tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag taaaacctct acaaatgtgg
     4441 taaaatcgat aaggatccag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     4501 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     4561 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     4621 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     4681 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     4741 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     4801 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     4861 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     4921 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     4981 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     5041 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     5101 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     5161 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     5221 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     5281 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     5341 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     5401 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     5461 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     5521 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     5581 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     5641 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     5701 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     5761 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     5821 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     5881 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     5941 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     6001 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     6061 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     6121 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     6181 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     6241 ggccttttgc tggccttttg ctcacatggc t
//
