LOCUS       sequence-214962-        6648 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3'UTR reporter assay.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6648)
  AUTHORS   Zhou Q, Anderson C, Hanus J, Zhao F, Ma J, Yoshimura A, Wang S
  TITLE     Strand and Cell Type-specific Function of microRNA-126 in 
            Angiogenesis.
  JOURNAL   Mol Ther. 2016 Oct;24(10):1823-1835. doi: 10.1038/mt.2016.108. Epub 
            2016 May 19.
  PUBMED    27203443
REFERENCE   2  (bases 1 to 6648)
  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: 
            "10.1038/mt.2016.108"; journalName: "Mol Ther"; date: "2016-10"; 
            volume: "24"; issue: "10"; pages: "1823-1835"
COMMENT     Sequence Label: pmiR-report-hVEGFA
FEATURES             Location/Qualifiers
     source          1..6648
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        76..379
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        380..583
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     534..554
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             608..2257
                     /codon_start=1
                     /gene="luc"
                     /product="firefly luciferase"
                     /label=luciferase
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVNITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMNISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIVPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKSKL"
     primer_bind     complement(674..692)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     primer_bind     complement(2591..2608)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2591..2607)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2600..2622)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2816..2835)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        2997..3313
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3164..3299
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3410..4009
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3410..3429)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3906..3926
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4239..4320
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4276..4295)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        4408..4512
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4513..4581,4582..5373)
                     /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(4731..4750)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5544..6132
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6033..6052
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6286..6303
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6420..6441
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6456..6461,6462..6479,6480..6486)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6494..6510
                     /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)."
     primer_bind     6499..6521
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6518..6534
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6518..6534
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        6554..6572
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     6554..6571
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     6584..6603
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6584..6602
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 caaggtacca gatcttagtt attaatagta atcaattacg gggtcattag ttcatagccc
       61 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
      121 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
      181 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
      241 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
      301 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
      361 agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg
      421 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
      481 gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat
      541 gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctggtttag tgaaccgtgg
      601 atccaccatg gaagacgcca aaaacataaa gaaaggcccg gcgccattct atcctctaga
      661 ggatggaacc gctggagagc aactgcataa ggctatgaag agatacgccc tggttcctgg
      721 aacaattgct tttacagatg cacatatcga ggtgaacatc acgtacgcgg aatacttcga
      781 aatgtccgtt cggttggcag aagctatgaa acgatatggg ctgaatacaa atcacagaat
      841 cgtcgtatgc agtgaaaact ctcttcaatt ctttatgccg gtgttgggcg cgttatttat
      901 cggagttgca gttgcgcccg cgaacgacat ttataatgaa cgtgaattgc tcaacagtat
      961 gaacatttcg cagcctaccg tagtgtttgt ttccaaaaag gggttgcaaa aaattttgaa
     1021 cgtgcaaaaa aaattaccaa taatccagaa aattattatc atggattcta aaacggatta
     1081 ccagggattt cagtcgatgt acacgttcgt cacatctcat ctacctcccg gttttaatga
     1141 atacgatttt gtaccagagt cctttgatcg tgacaaaaca attgcactga taatgaattc
     1201 ctctggatct actgggttac ctaagggtgt ggcccttccg catagaactg cctgcgtcag
     1261 attctcgcat gccagagatc ctatttttgg caatcaaatc gttccggata ctgcgatttt
     1321 aagtgttgtc ccattccatc acggttttgg aatgtttact acactcggat atttgatatg
     1381 tggatttcga gtcgtcttaa tgtatagatt tgaagaagag ctgtttttac gatcccttca
     1441 ggattacaaa attcaaagtg cgttgctagt accaacccta ttttcattct tcgccaaaag
     1501 cactctgatt gacaaatacg atttatctaa tttacacgaa attgcttctg ggggcgcacc
     1561 tctttcgaaa gaagtcgggg aagcggttgc aaaacgcttc catcttccag ggatacgaca
     1621 aggatatggg ctcactgaga ctacatcagc tattctgatt acacccgagg gggatgataa
     1681 accgggcgcg gtcggtaaag ttgttccatt ttttgaagcg aaggttgtgg atctggatac
     1741 cgggaaaacg ctgggcgtta atcagagagg cgaattatgt gtcagaggac ctatgattat
     1801 gtccggttat gtaaacaatc cggaagcgac caacgccttg attgacaagg atggatggct
     1861 acattctgga gacatagctt actgggacga agacgaacac ttcttcatag ttgaccgctt
     1921 gaagtcttta attaaataca aaggatatca ggtggccccc gctgaattgg aatcgatatt
     1981 gttacaacac cccaacatct tcgacgcggg cgtggcaggt cttcccgacg atgacgccgg
     2041 tgaacttccc gccgccgttg ttgttttgga gcacggaaag acgatgacgg aaaaagagat
     2101 cgtggattac gtcgccagtc aagtaacaac cgcgaaaaag ttgcgcggag gagttgtgtt
     2161 tgtggacgaa gtaccgaaag gtcttaccgg aaaactcgac gcaagaaaaa tcagagagat
     2221 cctcataaag gccaagaagg gcggaaagtc caaattgctc gagtgatgaa agctgcgcac
     2281 tagtgaggga gctcccttcc tcatcttccc ttcccaaggc acagagcaat ggggcaggac
     2341 ccgcaagccc ctcacggagg cagagaaaag agaaagtgtt ttatatacgg tacttattta
     2401 atagcccttt ttaattagaa attaaaacag ttaatttaat taaagagtag ggtttttttc
     2461 agtattcttg gttaatattt aatttcaact atttatgaga tgtatctctc gctctctcaa
     2521 gcttaataaa ggatctttta ttttcattgg atctgtgtgt tggttttttg tatgcggccg
     2581 ctagcttggc actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc
     2641 aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc
     2701 gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatggcgc ctgatgcggt
     2761 attttctcct tacgcatctg tgcggtattt cacaccgcat atggtgcact ctcagtacaa
     2821 tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc
     2881 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga
     2941 gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagattggtg
     3001 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     3061 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     3121 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     3181 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     3241 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     3301 aggcttttgc aaaaagctag cttgcatgcc tgcaggtcgg ccgccacgac cggtgccgcc
     3361 accatcccct gacccacgcc cctgacccct cacaaggaga cgaccttcca tgaccgagta
     3421 caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc cgggccgtac gcaccctcgc
     3481 cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc gacccggacc gccacatcga
     3541 gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt
     3601 gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc
     3661 gggggcggtg ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc
     3721 cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt
     3781 cctggccacc gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt
     3841 gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacctccgc
     3901 gccccgcaac ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt
     3961 gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgac gcccgcccca
     4021 cgacccgcag cgcccgaccg aaaggagcgc acgaccccat ggctccgacc gaagccaccc
     4081 ggggcggccc cgccgacccc gcacccgccc ccgaggccca ccgactctag aggatcataa
     4141 tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc
     4201 tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata
     4261 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     4321 aatctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg
     4381 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     4441 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     4501 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     4561 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     4621 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     4681 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     4741 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     4801 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     4861 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     4921 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     4981 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     5041 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     5101 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     5161 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     5221 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     5281 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     5341 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     5401 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     5461 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     5521 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     5581 caaacaaaaa aaccaccgct accagcggtg gtttttttgc cggatcaaga gctaccaact
     5641 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     5701 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     5761 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     5821 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     5881 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     5941 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     6001 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     6061 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     6121 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     6181 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     6241 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     6301 gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat
     6361 taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt
     6421 aatgtgagtt agctcactca ttaggcaccc caggctttac actttatgct tccggctcgt
     6481 atgttgtgtg gaattgtgag cggataacaa tttcacacag gaaacagcta tgacatgatt
     6541 acgaattgca acgatttagg tgacactata gaagagaagg aattaatacg actcactata
     6601 gggagagaga gagaattacc ctcactaaag ggaggagaag catgaatt
//
