LOCUS       sequence-322211-        6270 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression vector, Luciferase reporter to test activity of
            miRNA (miR-497).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6270)
  TITLE     Karreth lab plasmids
REFERENCE   2  (bases 1 to 6270)
  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-497
FEATURES             Location/Qualifiers
     source          1..6270
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        62..419
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        62..253
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(85..105)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      270..405
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          489..621
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     604..623
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        666..684
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      688..697
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             694..1629
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     polyA_signal    1685..1733
                     /note="synthetic polyadenylation signal"
     promoter        1742..2493
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     regulatory      2523..2532
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2529..4181
                     /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             3488..3496
                     /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    4225..4346
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4236..4255)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4290..4309
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        4479..4583
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4584..4652,4653..5444)
                     /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(4802..4821)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5615..6203
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6104..6123
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agatctgcgc agcaccatgg cctgaaataa cctctgaaag aggaacttgg ttaggtacct
       61 tctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
      121 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
      181 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
      241 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
      301 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg
      361 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
      421 agcttgattc ttctgacaca acagtctcga acttaagctg cagaagttgg tcgtgaggca
      481 ctgggcaggt aagtatcaag gttacaagac aggtttaagg agaccaatag aaactgggct
      541 tgtcgagaca gagaagactc ttgcgtttct gataggcacc tattggtctt actgacatcc
      601 actttgcctt tctctccaca ggtgtccact cccagttcaa ttacagctct taaggctaga
      661 gtacttaata cgactcacta taggctagcc accatggctt ccaaggtgta cgaccccgag
      721 caacgcaaac gcatgatcac tgggcctcag tggtgggctc gctgcaagca aatgaacgtg
      781 ctggactcct tcatcaacta ctatgattcc gagaagcacg ccgagaacgc cgtgattttt
      841 ctgcatggta acgctgcctc cagctacctg tggaggcacg tcgtgcctca catcgagccc
      901 gtggctagat gcatcatccc tgatctgatc ggaatgggta agtccggcaa gagcgggaat
      961 ggctcatatc gcctcctgga tcactacaag tacctcaccg cttggttcga gctgctgaac
     1021 cttccaaaga aaatcatctt tgtgggccac gactgggggg cttgtctggc ctttcactac
     1081 tcctacgagc accaagacaa gatcaaggcc atcgtccatg ctgagagtgt cgtggacgtg
     1141 atcgagtcct gggacgagtg gcctgacatc gaggaggata tcgccctgat caagagcgaa
     1201 gagggcgaga aaatggtgct tgagaataac ttcttcgtcg agaccatgct cccaagcaag
     1261 atcatgcgga aactggagcc tgaggagttc gctgcctacc tggagccatt caaggagaag
     1321 ggcgaggtta gacggcctac cctctcctgg cctcgcgaga tccctctcgt taagggaggc
     1381 aagcccgacg tcgtccagat tgtccgcaac tacaacgcct accttcgggc cagcgacgat
     1441 ctgcctaaga tgttcatcga gtccgaccct gggttctttt ccaacgctat tgtcgaggga
     1501 gctaagaagt tccctaacac cgagttcgtg aaggtgaagg gcctccactt cagccaggag
     1561 gacgctccag atgaaatggg taagtacatc aagagcttcg tggagcgcgt gctgaagaac
     1621 gagcagtaat tctaggcgat cgctcgacac aaaccacagt gtgctgctgg cggccgctgg
     1681 ccgcaataaa atatctttat tttcattaca tctgtgtgtt ggttttttgt gtgaggatct
     1741 aaatgagtct tcggacctcg cgggggccgc ttaagcggtg gttagggttt gtctgacgcg
     1801 gggggagggg gaaggaacga aacactctca ttcggaggcg gctcggggtt tggtcttggt
     1861 ggccacgggc acgcagaaga gcgccgcgat cctcttaagc acccccccgc cctccgtgga
     1921 ggcgggggtt tggtcggcgg gtggtaactg gcgggccgct gactcgggcg ggtcgcgcgc
     1981 cccagagtgt gaccttttcg gtctgctcgc agacccccgg gcggcgccgc cgcggcggcg
     2041 acgggctcgc tgggtcctag gctccatggg gaccgtatac gtggacaggc tctggagcat
     2101 ccgcacgact gcggtgatat taccggagac cttctgcggg acgagccggg tcacgcggct
     2161 gacgcggagc gtccgttggg cgacaaacac caggacgggg cacaggtaca ctatcttgtc
     2221 acccggaggc gcgagggact gcaggagctt cagggagtgg cgcagctgct tcatccccgt
     2281 ggcccgttgc tcgcgtttgc tggcggtgtc cccggaagaa atatatttgc atgtctttag
     2341 ttctatgatg acacaaaccc cgcccagcgt cttgtcattg gcgaattcga acacgcagat
     2401 gcagtcgggg cggcgcggtc ccaggtccac ttcgcatatt aaggtgacgc gtgtggcctc
     2461 gaacaccgag cgaccctgca gcgacccgct taaaagcttg gcattccggt actgttggta
     2521 aagccaccat ggccgatgct aagaacatta agaagggccc tgctcccttc taccctctgg
     2581 aggatggcac cgctggcgag cagctgcaca aggccatgaa gaggtatgcc ctggtgcctg
     2641 gcaccattgc cttcaccgat gcccacattg aggtggacat cacctatgcc gagtacttcg
     2701 agatgtctgt gcgcctggcc gaggccatga agaggtacgg cctgaacacc aaccaccgca
     2761 tcgtggtgtg ctctgagaac tctctgcagt tcttcatgcc agtgctgggc gccctgttca
     2821 tcggagtggc cgtggcccct gctaacgaca tttacaacga gcgcgagctg ctgaacagca
     2881 tgggcatttc tcagcctacc gtggtgttcg tgtctaagaa gggcctgcag aagatcctga
     2941 acgtgcagaa gaagctgcct atcatccaga agatcatcat catggactct aagaccgact
     3001 accagggctt ccagagcatg tacacattcg tgacatctca tctgcctcct ggcttcaacg
     3061 agtacgactt cgtgccagag tctttcgaca gggacaaaac cattgccctg atcatgaaca
     3121 gctctgggtc taccggcctg cctaagggcg tggccctgcc tcatcgcacc gcctgtgtgc
     3181 gcttctctca cgcccgcgac cctattttcg gcaaccagat catccccgac accgctattc
     3241 tgagcgtggt gccattccac cacggcttcg gcatgttcac caccctgggc tacctgattt
     3301 gcggctttcg ggtggtgctg atgtaccgct tcgaggagga gctgttcctg cgcagcctgc
     3361 aagactacaa aattcagtct gccctgctgg tgccaaccct gttcagcttc ttcgctaaga
     3421 gcaccctgat cgacaagtac gacctgtcta acctgcacga gattgcctct ggcggcgccc
     3481 cactgtctaa ggaggtgggc gaagccgtgg ccaagcgctt tcatctgcca ggcatccgcc
     3541 agggctacgg cctgaccgag acaaccagcg ccattctgat taccccagag ggcgacgaca
     3601 agcctggcgc cgtgggcaag gtggtgccat tcttcgaggc caaggtggtg gacctggaca
     3661 ccggcaagac cctgggagtg aaccagcgcg gcgagctgtg tgtgcgcggc cctatgatta
     3721 tgtccggcta cgtgaataac cctgaggcca caaacgccct gatcgacaag gacggctggc
     3781 tgcactctgg cgacattgcc tactgggacg aggacgagca cttcttcatc gtggaccgcc
     3841 tgaagtctct gatcaagtac aagggctacc aggtggcccc agccgagctg gagtctatcc
     3901 tgctgcagca ccctaacatt ttcgacgccg gagtggccgg cctgcccgac gacgatgccg
     3961 gcgagctgcc tgccgccgtc gtcgtgctgg aacacggcaa gaccatgacc gagaaggaga
     4021 tcgtggacta tgtggccagc caggtgacaa ccgccaagaa gctgcgcggc ggagtggtgt
     4081 tcgtggacga ggtgcccaag ggcctgaccg gcaagctgga cgcccgcaag atccgcgaga
     4141 tcctgatcaa ggctaagaaa ggcggcaaga tcgccgtgta ataattctag agtcggggcg
     4201 gccggccgct tcgagcagac atgataagat acattgatga gtttggacaa accacaacta
     4261 gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga tgctattgct ttatttgtaa
     4321 ccattataag ctgcaataaa caagttaaca acaacaattg cattcatttt atgtttcagg
     4381 ttcaggggga ggtgtgggag gttttttaaa gcaagtaaaa cctctacaaa tgtggtaaaa
     4441 tcgataagga tccaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     4501 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     4561 aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct
     4621 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     4681 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta
     4741 agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc
     4801 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     4861 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg
     4921 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     4981 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca
     5041 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     5101 acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa
     5161 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata
     5221 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     5281 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     5341 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     5401 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt
     5461 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     5521 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     5581 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     5641 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     5701 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     5761 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     5821 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     5881 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     5941 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     6001 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     6061 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     6121 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     6181 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     6241 tttgctggcc ttttgctcac atggctcgac
//
