LOCUS       sequence-73907-e        4435 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Promoterless luc2 donor plasmid for MPRA library construction.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4435)
  AUTHORS   Melnikov A, Zhang X, Rogov P, Wang L, Mikkelsen TS
  TITLE     Massively parallel reporter assays in cultured Mammalian cells.
  JOURNAL   J Vis Exp. 2014 Aug 17;(90). doi: 10.3791/51719.
  PUBMED    25177895
REFERENCE   2  (bases 1 to 4435)
  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: "J Vis 
            Exp."; date: "2014-08-17"; volume: "(90). doi"; pages: " 
            10.3791/51719"
COMMENT     Sequence Label: pMPRAdonor1
FEATURES             Location/Qualifiers
     source          1..4435
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      81..90
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1807..1823)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1807..1823)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1820..1842)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1831..1847
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(1855..1861,1862..1879,1880..1885))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1900..1921
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2038..2055)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2209..2797)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2289..2308)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2968..3759,3760..3828))
                     /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     3591..3610
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3829..3933)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4001..4019
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4057..4079)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4179..4198
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4392..4414
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4406..4423
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4407..4423
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 ggtacctgag ctcgctagcc tcgaggatat caagatctgg cctcggcggc caagcttggc
       61 aatccggtac tgttggtaaa gccaccatgg aagatgccaa aaacattaag aagggcccag
      121 cgccattcta cccactcgaa gacgggaccg ccggcgagca gctgcacaaa gccatgaagc
      181 gctacgccct ggtgcccggc accatcgcct ttaccgacgc acatatcgag gtggacatta
      241 cctacgccga gtacttcgag atgagcgttc ggctggcaga agctatgaag cgctatgggc
      301 tgaatacaaa ccatcggatc gtggtgtgca gcgagaatag cttgcagttc ttcatgcccg
      361 tgttgggtgc cctgttcatc ggtgtggctg tggccccagc taacgacatc tacaacgagc
      421 gcgagctgct gaacagcatg ggcatcagcc agcccaccgt cgtattcgtg agcaagaaag
      481 ggctgcaaaa gatcctcaac gtgcaaaaga agctaccgat catacaaaag atcatcatca
      541 tggatagcaa gaccgactac cagggcttcc aaagcatgta caccttcgtg acttcccatt
      601 tgccacccgg cttcaacgag tacgacttcg tgcccgagag cttcgaccgg gacaaaacca
      661 tcgccctgat catgaacagt agtggcagta ccggattgcc caagggcgta gccctaccgc
      721 accgcaccgc ttgtgtccga ttcagtcatg cccgcgaccc catcttcggc aaccagatca
      781 tccccgacac cgctatcctc agcgtggtgc catttcacca cggcttcggc atgttcacca
      841 cgctgggcta cttgatctgc ggctttcggg tcgtgctcat gtaccgcttc gaggaggagc
      901 tattcttgcg cagcttgcaa gactataaga ttcaatctgc cctgctggtg cccacactat
      961 ttagcttctt cgctaagagc actctcatcg acaagtacga cctaagcaac ttgcacgaga
     1021 tcgccagcgg cggggcgccg ctcagcaagg aggtaggtga ggccgtggcc aaacgcttcc
     1081 acctaccagg catccgccag ggctacggcc tgacagaaac aaccagcgcc attctgatca
     1141 cccccgaagg ggacgacaag cctggcgcag taggcaaggt ggtgcccttc ttcgaggcta
     1201 aggtggtgga cttggacacc ggtaagacac tgggtgtgaa ccagcgcggc gagctgtgcg
     1261 tccgtggccc catgatcatg acggctacgt taacaacccc gaggctacaa acgctctcat
     1321 cgacaaggac ggctggctgc acagcggcga catcgcctac tgggacgagg acgagcactt
     1381 cttcatcgtg gaccggctga agagcctgat caaatacaag ggctaccagg tagccccagc
     1441 cgaactggag agcatcctgc tgcaacaccc caacatcttc gacgccgggg tcgccggcct
     1501 gcccgacgac gatgccggcg agctgcccgc cgcagtcgtc gtgctggaac acggtaaaac
     1561 catgaccgag aaggagatcg tggactatgt ggccagccag gttacaaccg ccaagaagct
     1621 gcgcggtggt gttgtgttcg tggacgaggt gcctaaagga ctgaccggca agttggacgc
     1681 ccgcaagatc cgcgagattc tcattaaggc caagaagggc ggcaagatcg ccgtgtaata
     1741 attctagata tcggatcccg ggcccgtcga ctgcagaggc ctgcatgcaa gcttggcgta
     1801 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     1861 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     1921 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     1981 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     2041 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     2101 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     2161 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     2221 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     2281 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     2341 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     2401 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     2461 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     2521 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     2581 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     2641 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     2701 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     2761 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     2821 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     2881 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     2941 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     3001 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     3061 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     3121 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     3181 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     3241 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     3301 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     3361 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     3421 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     3481 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     3541 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     3601 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     3661 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     3721 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     3781 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     3841 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     3901 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     3961 cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc
     4021 ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga
     4081 gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc
     4141 agcgggtgtt ggcgggtgtc ggggctggct taactatgcg gcatcagagc agattgtact
     4201 gagagtgcac catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat
     4261 caggcgccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc
     4321 ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac
     4381 gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgaattcg agctc
//
