LOCUS       sequence-307133-        7160 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7160)
  AUTHORS   Melton C, Judson RL, Blelloch R
  TITLE     Opposing microRNA families regulate self-renewal in mouse embryonic 
            stem cells.
  JOURNAL   Nature. 2010 Feb 4;463(7281):621-6. Epub 2010 Jan 6.
  PUBMED    20054295
REFERENCE   2  (bases 1 to 7160)
  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: "Nature."; 
            date: "2010-02-4"; volume: "463(7281)"; pages: "621-6. Epub 2010 Jan
            6"
COMMENT     Sequence Label: psiCHECK-2-nMyc-wt_3'UTR
FEATURES             Location/Qualifiers
     source          1..7160
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        83..440
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        83..274
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(106..126)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      291..426
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          510..642
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     625..644
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        687..705
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      709..718
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             715..1650
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     polyA_signal    2596..2644
                     /note="synthetic polyadenylation signal"
     promoter        2653..3404
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     regulatory      3434..3443
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3440..5092
                     /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             4399..4407
                     /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    5136..5257
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5147..5166)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5201..5220
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        5390..5494
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5495..5563,5564..6355)
                     /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(5713..5732)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6526..7114
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7015..7034
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cttttgctca catggctcga cagatctgcg cagcaccatg gcctgaaata acctctgaaa
       61 gaggaacttg gttaggtacc ttctgaggcg gaaagaacca gctgtggaat gtgtgtcagt
      121 tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
      181 attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
      241 gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc
      301 taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg
      361 cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg
      421 gaggcctagg cttttgcaaa aagcttgatt cttctgacac aacagtctcg aacttaagct
      481 gcagaagttg gtcgtgaggc actgggcagg taagtatcaa ggttacaaga caggtttaag
      541 gagaccaata gaaactgggc ttgtcgagac agagaagact cttgcgtttc tgataggcac
      601 ctattggtct tactgacatc cactttgcct ttctctccac aggtgtccac tcccagttca
      661 attacagctc ttaaggctag agtacttaat acgactcact ataggctagc caccatggct
      721 tccaaggtgt acgaccccga gcaacgcaaa cgcatgatca ctgggcctca gtggtgggct
      781 cgctgcaagc aaatgaacgt gctggactcc ttcatcaact actatgattc cgagaagcac
      841 gccgagaacg ccgtgatttt tctgcatggt aacgctgcct ccagctacct gtggaggcac
      901 gtcgtgcctc acatcgagcc cgtggctaga tgcatcatcc ctgatctgat cggaatgggt
      961 aagtccggca agagcgggaa tggctcatat cgcctcctgg atcactacaa gtacctcacc
     1021 gcttggttcg agctgctgaa ccttccaaag aaaatcatct ttgtgggcca cgactggggg
     1081 gcttgtctgg cctttcacta ctcctacgag caccaagaca agatcaaggc catcgtccat
     1141 gctgagagtg tcgtggacgt gatcgagtcc tgggacgagt ggcctgacat cgaggaggat
     1201 atcgccctga tcaagagcga agagggcgag aaaatggtgc ttgagaataa cttcttcgtc
     1261 gagaccatgc tcccaagcaa gatcatgcgg aaactggagc ctgaggagtt cgctgcctac
     1321 ctggagccat tcaaggagaa gggcgaggtt agacggccta ccctctcctg gcctcgcgag
     1381 atccctctcg ttaagggagg caagcccgac gtcgtccaga ttgtccgcaa ctacaacgcc
     1441 taccttcggg ccagcgacga tctgcctaag atgttcatcg agtccgaccc tgggttcttt
     1501 tccaacgcta ttgtcgaggg agctaagaag ttccctaaca ccgagttcgt gaaggtgaag
     1561 ggcctccact tcagccagga ggacgctcca gatgaaatgg gtaagtacat caagagcttc
     1621 gtggagcgcg tgctgaagaa cgagcagtaa ttctaggcga tcgctcgagc gtttcccaca
     1681 cggacagtca ctgccacttt gcacattttg attttttttt tttttttaaa caaacattgt
     1741 gttgacatta agaatgttgg tttactttca aattggtccc ctgtcgagtc tggatctggg
     1801 tagggggcag gacacggggt tctgccatga ccttggaaaa aaaactgact tatgggatgc
     1861 tgggtggctt gttttcctcc tccatatcac ctggtgacag ccgtggaagt tcgggacact
     1921 aaggagcttc aggaggctgt gaagtcacct tgttccggtc caagattcca aacagagtca
     1981 ttccttcttt ttacaatggt gcttaagttc cagcaaatgc cacagaaggg ggggttgcca
     2041 tttgatgccc ctgggaacac ttgtgtaaat accattgata cgcccgcctt ttgtatacgt
     2101 cctgggtaat gagaggtggc tcttgcggcc agtattagac tggaagttca cacctaagta
     2161 ctgtaagaat acctcaatgt ttgaggggca tgttttgtat acaaatatat tgttaatctg
     2221 ttatgtactg tactaattcc tacacggcct atatacttta gtatgacgct gatacataac
     2281 taaatttgat acttatattt tcgtatgaaa atgagttgtg gaagttttga gtagatatta
     2341 ctttatcact ttttgaacta agaaactttt gtaaagaaat tttactatat atatatattc
     2401 ctttttttcc tagcctgttt cttccttgtt tactgtattt gttcatgttt ggtgcataga
     2461 actgggtaaa atggcaaagt tctgtgttta atttcttcaa aatgtatata tttagtgctg
     2521 caccttagag cactttgaaa tacctcatgt ttatgaaaat aaatagcaat taaatgatgc
     2581 gcggccgctg gccgcaataa aatatcttta ttttcattac atctgtgtgt tggttttttg
     2641 tgtgaggatc taaatgagtc ttcggacctc gcgggggccg cttaagcggt ggttagggtt
     2701 tgtctgacgc ggggggaggg ggaaggaacg aaacactctc attcggaggc ggctcggggt
     2761 ttggtcttgg tggccacggg cacgcagaag agcgccgcga tcctcttaag cacccccccg
     2821 ccctccgtgg aggcgggggt ttggtcggcg ggtggtaact ggcgggccgc tgactcgggc
     2881 gggtcgcgcg ccccagagtg tgaccttttc ggtctgctcg cagacccccg ggcggcgccg
     2941 ccgcggcggc gacgggctcg ctgggtccta ggctccatgg ggaccgtata cgtggacagg
     3001 ctctggagca tccgcacgac tgcggtgata ttaccggaga ccttctgcgg gacgagccgg
     3061 gtcacgcggc tgacgcggag cgtccgttgg gcgacaaaca ccaggacggg gcacaggtac
     3121 actatcttgt cacccggagg cgcgagggac tgcaggagct tcagggagtg gcgcagctgc
     3181 ttcatccccg tggcccgttg ctcgcgtttg ctggcggtgt ccccggaaga aatatatttg
     3241 catgtcttta gttctatgat gacacaaacc ccgcccagcg tcttgtcatt ggcgaattcg
     3301 aacacgcaga tgcagtcggg gcggcgcggt cccaggtcca cttcgcatat taaggtgacg
     3361 cgtgtggcct cgaacaccga gcgaccctgc agcgacccgc ttaaaagctt ggcattccgg
     3421 tactgttggt aaagccacca tggccgatgc taagaacatt aagaagggcc ctgctccctt
     3481 ctaccctctg gaggatggca ccgctggcga gcagctgcac aaggccatga agaggtatgc
     3541 cctggtgcct ggcaccattg ccttcaccga tgcccacatt gaggtggaca tcacctatgc
     3601 cgagtacttc gagatgtctg tgcgcctggc cgaggccatg aagaggtacg gcctgaacac
     3661 caaccaccgc atcgtggtgt gctctgagaa ctctctgcag ttcttcatgc cagtgctggg
     3721 cgccctgttc atcggagtgg ccgtggcccc tgctaacgac atttacaacg agcgcgagct
     3781 gctgaacagc atgggcattt ctcagcctac cgtggtgttc gtgtctaaga agggcctgca
     3841 gaagatcctg aacgtgcaga agaagctgcc tatcatccag aagatcatca tcatggactc
     3901 taagaccgac taccagggct tccagagcat gtacacattc gtgacatctc atctgcctcc
     3961 tggcttcaac gagtacgact tcgtgccaga gtctttcgac agggacaaaa ccattgccct
     4021 gatcatgaac agctctgggt ctaccggcct gcctaagggc gtggccctgc ctcatcgcac
     4081 cgcctgtgtg cgcttctctc acgcccgcga ccctattttc ggcaaccaga tcatccccga
     4141 caccgctatt ctgagcgtgg tgccattcca ccacggcttc ggcatgttca ccaccctggg
     4201 ctacctgatt tgcggctttc gggtggtgct gatgtaccgc ttcgaggagg agctgttcct
     4261 gcgcagcctg caagactaca aaattcagtc tgccctgctg gtgccaaccc tgttcagctt
     4321 cttcgctaag agcaccctga tcgacaagta cgacctgtct aacctgcacg agattgcctc
     4381 tggcggcgcc ccactgtcta aggaggtggg cgaagccgtg gccaagcgct ttcatctgcc
     4441 aggcatccgc cagggctacg gcctgaccga gacaaccagc gccattctga ttaccccaga
     4501 gggcgacgac aagcctggcg ccgtgggcaa ggtggtgcca ttcttcgagg ccaaggtggt
     4561 ggacctggac accggcaaga ccctgggagt gaaccagcgc ggcgagctgt gtgtgcgcgg
     4621 ccctatgatt atgtccggct acgtgaataa ccctgaggcc acaaacgccc tgatcgacaa
     4681 ggacggctgg ctgcactctg gcgacattgc ctactgggac gaggacgagc acttcttcat
     4741 cgtggaccgc ctgaagtctc tgatcaagta caagggctac caggtggccc cagccgagct
     4801 ggagtctatc ctgctgcagc accctaacat tttcgacgcc ggagtggccg gcctgcccga
     4861 cgacgatgcc ggcgagctgc ctgccgccgt cgtcgtgctg gaacacggca agaccatgac
     4921 cgagaaggag atcgtggact atgtggccag ccaggtgaca accgccaaga agctgcgcgg
     4981 cggagtggtg ttcgtggacg aggtgcccaa gggcctgacc ggcaagctgg acgcccgcaa
     5041 gatccgcgag atcctgatca aggctaagaa aggcggcaag atcgccgtgt aataattcta
     5101 gagtcggggc ggccggccgc ttcgagcaga catgataaga tacattgatg agtttggaca
     5161 aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc
     5221 tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt gcattcattt
     5281 tatgtttcag gttcaggggg aggtgtggga ggttttttaa agcaagtaaa acctctacaa
     5341 atgtggtaaa atcgataagg atccaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     5401 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     5461 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     5521 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     5581 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     5641 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     5701 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact
     5761 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     5821 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     5881 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     5941 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     6001 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg
     6061 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     6121 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     6181 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc
     6241 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     6301 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     6361 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     6421 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     6481 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     6541 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     6601 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     6661 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     6721 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     6781 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     6841 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     6901 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     6961 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     7021 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     7081 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     7141 gttcctggcc ttttgctggc
//
