LOCUS       sequence-75541-e        3824 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Template DNA for in vitro transcription using T7 polymerase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3824)
  AUTHORS   Fukaya T, Tomari Y
  TITLE     PABP is not essential for microRNA-mediated translational repression
            and deadenylation in vitro.
  JOURNAL   EMBO J. 2011 Nov 25;30(24):4998-5009. doi: 10.1038/emboj.2011.426.
  PUBMED    22117217
REFERENCE   2  (bases 1 to 3824)
  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/emboj.2011"; journalName: "EMBO J"; date: "2011-11-25- 25";
            volume: "30"; issue: "24"; pages: "4998-5009"
COMMENT     Sequence Label: pUC57-Rluc-A114-N40
FEATURES             Location/Qualifiers
     source          1..3824
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        408..426
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      430..439
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             436..1371
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     primer_bind     complement(1603..1619)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1603..1619)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1616..1638)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1627..1643
                     /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(1651..1657,1658..1675,1676..1681))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1696..1717
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1834..1851)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2005..2593)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2085..2104)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2764..3555,3556..3624))
                     /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     3387..3406
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3625..3729)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3797..3815
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt cgagctctaa tacgactcac
      421 tataggctag ccaccatggc ttccaaggtg tacgaccccg agcaacgcaa acgcatgatc
      481 actgggcctc agtggtgggc tcgctgcaag caaatgaacg tgctggactc cttcatcaac
      541 tactatgatt ccgagaagca cgccgagaac gccgtgattt ttctgcatgg taacgctgcc
      601 tccagctacc tgtggaggca cgtcgtgcct cacatcgagc ccgtggctag atgcatcatc
      661 cctgatctga tcggaatggg taagtccggc aagagcggga atggctcata tcgcctcctg
      721 gatcactaca agtacctcac cgcttggttc gagctgctga accttccaaa gaaaatcatc
      781 tttgtgggcc acgactgggg ggcttgtctg gcctttcact actcctacga gcaccaagac
      841 aagatcaagg ccatcgtcca tgctgagagt gtcgtggacg tgatcgagtc ctgggacgag
      901 tggcctgaca tcgaggagga tatcgccctg atcaagagcg aagagggcga gaaaatggtg
      961 cttgagaata acttcttcgt cgagaccatg ctcccaagca agatcatgcg gaaactggag
     1021 cctgaggagt tcgctgccta cctggagcca ttcaaggaga agggcgaggt tagacggcct
     1081 accctctcct ggcctcgcga gatccctctc gttaagggag gcaagcccga cgtcgtccag
     1141 attgtccgca actacaacgc ctaccttcgg gccagcgacg atctgcctaa gatgttcatc
     1201 gagtccgacc ctgggttctt ttccaacgct attgtcgagg gagctaagaa gttccctaac
     1261 accgagttcg tgaaggtgaa gggcctccac ttcagccagg aggacgctcc agatgaaatg
     1321 ggtaagtaca tcaagagctt cgtggagcgc gtgctgaaga acgagcagta attctaggcg
     1381 atcgctcgag cccgggaatt cgtttaaacc tagagcggcc gctggccgcc tgcagaaaaa
     1441 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa
     1501 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaat gcatctggtt
     1561 tcctggtctg tctcctctcg gtttaagctt ggcgtaatca tggtcatagc tgtttcctgt
     1621 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     1681 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     1741 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     1801 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     1861 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     1921 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     1981 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     2041 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     2101 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     2161 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     2221 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     2281 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     2341 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     2401 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     2461 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     2521 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     2581 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     2641 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     2701 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     2761 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     2821 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     2881 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     2941 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     3001 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     3061 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     3121 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     3181 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     3241 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     3301 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     3361 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     3421 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     3481 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     3541 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     3601 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     3661 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     3721 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat
     3781 gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtc
//
