LOCUS       sequence-307132-        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-m2_3'UTR
FEATURES             Location/Qualifiers
     source          1..7160
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        61..418
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        61..252
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(84..104)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      269..404
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          488..620
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     603..622
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        665..683
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      687..696
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             693..1628
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     polyA_signal    2574..2622
                     /note="synthetic polyadenylation signal"
     promoter        2631..3382
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     regulatory      3412..3421
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3418..5070
                     /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             4377..4385
                     /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    5114..5235
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5125..5144)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5179..5198
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        5368..5472
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5473..5541,5542..6333)
                     /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(5691..5710)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6504..7092
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6993..7012
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gatctgcgca gcaccatggc ctgaaataac ctctgaaaga ggaacttggt taggtacctt
       61 ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
      121 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
      181 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
      241 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
      301 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc
      361 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
      421 gcttgattct tctgacacaa cagtctcgaa cttaagctgc agaagttggt cgtgaggcac
      481 tgggcaggta agtatcaagg ttacaagaca ggtttaagga gaccaataga aactgggctt
      541 gtcgagacag agaagactct tgcgtttctg ataggcacct attggtctta ctgacatcca
      601 ctttgccttt ctctccacag gtgtccactc ccagttcaat tacagctctt aaggctagag
      661 tacttaatac gactcactat aggctagcca ccatggcttc caaggtgtac gaccccgagc
      721 aacgcaaacg catgatcact gggcctcagt ggtgggctcg ctgcaagcaa atgaacgtgc
      781 tggactcctt catcaactac tatgattccg agaagcacgc cgagaacgcc gtgatttttc
      841 tgcatggtaa cgctgcctcc agctacctgt ggaggcacgt cgtgcctcac atcgagcccg
      901 tggctagatg catcatccct gatctgatcg gaatgggtaa gtccggcaag agcgggaatg
      961 gctcatatcg cctcctggat cactacaagt acctcaccgc ttggttcgag ctgctgaacc
     1021 ttccaaagaa aatcatcttt gtgggccacg actggggggc ttgtctggcc tttcactact
     1081 cctacgagca ccaagacaag atcaaggcca tcgtccatgc tgagagtgtc gtggacgtga
     1141 tcgagtcctg ggacgagtgg cctgacatcg aggaggatat cgccctgatc aagagcgaag
     1201 agggcgagaa aatggtgctt gagaataact tcttcgtcga gaccatgctc ccaagcaaga
     1261 tcatgcggaa actggagcct gaggagttcg ctgcctacct ggagccattc aaggagaagg
     1321 gcgaggttag acggcctacc ctctcctggc ctcgcgagat ccctctcgtt aagggaggca
     1381 agcccgacgt cgtccagatt gtccgcaact acaacgccta ccttcgggcc agcgacgatc
     1441 tgcctaagat gttcatcgag tccgaccctg ggttcttttc caacgctatt gtcgagggag
     1501 ctaagaagtt ccctaacacc gagttcgtga aggtgaaggg cctccacttc agccaggagg
     1561 acgctccaga tgaaatgggt aagtacatca agagcttcgt ggagcgcgtg ctgaagaacg
     1621 agcagtaatt ctaggcgatc gctcgagcgt ttcccacacg gacagtcact gccactttgc
     1681 acattttgat tttttttttt tttttaaaca aacattgtgt tgacattaag aatgttggtt
     1741 tactttcaaa ttggtcccct gtcgagtctg gatctgggta gggggcagga cacggggttc
     1801 tgccatgacc ttggaaaaaa aactgactta tgggatgctg ggtggcttgt tttcctcctc
     1861 catatcacct ggtgacagcc gtggaagttc gggacactaa ggagcttcag gaggctgtga
     1921 agtcaccttg ttccggtcca agattccaaa cagagtcatt ccttcttttt acaatggtgc
     1981 ttaagttcca gcaaatgcca cagaaggggg ggttgccatt tgatgcccct gggaacactt
     2041 gtgtaaatac cattgatacg cccgcctttt gtatacgtcc tgggtaatga gaggtggctc
     2101 ttgcggccag tattagactg gaagttcaca cctaagtact gtaagaatac ctcaatgttt
     2161 gaggggcatg ttttgtatac aaatatattg ttaatctgtt atgtactgta ctaattccta
     2221 cacggcctat atactttagt atgacgctga tacataacta aatttgatac ttatattttc
     2281 gtatgaaaat gagttgtgga agttttgagt agatattact ttatcacttt ttgaactaag
     2341 aaacttttgt aaagaaattt tactatatat atatattcct ttttttccta gcctgtttct
     2401 tccttgttta ctgtatttgt tcatgtttgg tgcatagaac tgggtaaaat ggcaaagttc
     2461 tgtgtttaat ttcttcaaaa tgtatatatt tagtgctgca ccttagagca ctttgaaata
     2521 acgcatgttt atgaaaataa atagcaatta aatgatgcgc ggccgctggc cgcaataaaa
     2581 tatctttatt ttcattacat ctgtgtgttg gttttttgtg tgaggatcta aatgagtctt
     2641 cggacctcgc gggggccgct taagcggtgg ttagggtttg tctgacgcgg ggggaggggg
     2701 aaggaacgaa acactctcat tcggaggcgg ctcggggttt ggtcttggtg gccacgggca
     2761 cgcagaagag cgccgcgatc ctcttaagca cccccccgcc ctccgtggag gcgggggttt
     2821 ggtcggcggg tggtaactgg cgggccgctg actcgggcgg gtcgcgcgcc ccagagtgtg
     2881 accttttcgg tctgctcgca gacccccggg cggcgccgcc gcggcggcga cgggctcgct
     2941 gggtcctagg ctccatgggg accgtatacg tggacaggct ctggagcatc cgcacgactg
     3001 cggtgatatt accggagacc ttctgcggga cgagccgggt cacgcggctg acgcggagcg
     3061 tccgttgggc gacaaacacc aggacggggc acaggtacac tatcttgtca cccggaggcg
     3121 cgagggactg caggagcttc agggagtggc gcagctgctt catccccgtg gcccgttgct
     3181 cgcgtttgct ggcggtgtcc ccggaagaaa tatatttgca tgtctttagt tctatgatga
     3241 cacaaacccc gcccagcgtc ttgtcattgg cgaattcgaa cacgcagatg cagtcggggc
     3301 ggcgcggtcc caggtccact tcgcatatta aggtgacgcg tgtggcctcg aacaccgagc
     3361 gaccctgcag cgacccgctt aaaagcttgg cattccggta ctgttggtaa agccaccatg
     3421 gccgatgcta agaacattaa gaagggccct gctcccttct accctctgga ggatggcacc
     3481 gctggcgagc agctgcacaa ggccatgaag aggtatgccc tggtgcctgg caccattgcc
     3541 ttcaccgatg cccacattga ggtggacatc acctatgccg agtacttcga gatgtctgtg
     3601 cgcctggccg aggccatgaa gaggtacggc ctgaacacca accaccgcat cgtggtgtgc
     3661 tctgagaact ctctgcagtt cttcatgcca gtgctgggcg ccctgttcat cggagtggcc
     3721 gtggcccctg ctaacgacat ttacaacgag cgcgagctgc tgaacagcat gggcatttct
     3781 cagcctaccg tggtgttcgt gtctaagaag ggcctgcaga agatcctgaa cgtgcagaag
     3841 aagctgccta tcatccagaa gatcatcatc atggactcta agaccgacta ccagggcttc
     3901 cagagcatgt acacattcgt gacatctcat ctgcctcctg gcttcaacga gtacgacttc
     3961 gtgccagagt ctttcgacag ggacaaaacc attgccctga tcatgaacag ctctgggtct
     4021 accggcctgc ctaagggcgt ggccctgcct catcgcaccg cctgtgtgcg cttctctcac
     4081 gcccgcgacc ctattttcgg caaccagatc atccccgaca ccgctattct gagcgtggtg
     4141 ccattccacc acggcttcgg catgttcacc accctgggct acctgatttg cggctttcgg
     4201 gtggtgctga tgtaccgctt cgaggaggag ctgttcctgc gcagcctgca agactacaaa
     4261 attcagtctg ccctgctggt gccaaccctg ttcagcttct tcgctaagag caccctgatc
     4321 gacaagtacg acctgtctaa cctgcacgag attgcctctg gcggcgcccc actgtctaag
     4381 gaggtgggcg aagccgtggc caagcgcttt catctgccag gcatccgcca gggctacggc
     4441 ctgaccgaga caaccagcgc cattctgatt accccagagg gcgacgacaa gcctggcgcc
     4501 gtgggcaagg tggtgccatt cttcgaggcc aaggtggtgg acctggacac cggcaagacc
     4561 ctgggagtga accagcgcgg cgagctgtgt gtgcgcggcc ctatgattat gtccggctac
     4621 gtgaataacc ctgaggccac aaacgccctg atcgacaagg acggctggct gcactctggc
     4681 gacattgcct actgggacga ggacgagcac ttcttcatcg tggaccgcct gaagtctctg
     4741 atcaagtaca agggctacca ggtggcccca gccgagctgg agtctatcct gctgcagcac
     4801 cctaacattt tcgacgccgg agtggccggc ctgcccgacg acgatgccgg cgagctgcct
     4861 gccgccgtcg tcgtgctgga acacggcaag accatgaccg agaaggagat cgtggactat
     4921 gtggccagcc aggtgacaac cgccaagaag ctgcgcggcg gagtggtgtt cgtggacgag
     4981 gtgcccaagg gcctgaccgg caagctggac gcccgcaaga tccgcgagat cctgatcaag
     5041 gctaagaaag gcggcaagat cgccgtgtaa taattctaga gtcggggcgg ccggccgctt
     5101 cgagcagaca tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga
     5161 aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc
     5221 tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag
     5281 gtgtgggagg ttttttaaag caagtaaaac ctctacaaat gtggtaaaat cgataaggat
     5341 ccaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     5401 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     5461 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     5521 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     5581 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     5641 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     5701 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     5761 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     5821 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     5881 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     5941 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     6001 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     6061 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     6121 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     6181 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     6241 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     6301 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     6361 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     6421 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     6481 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     6541 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     6601 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     6661 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     6721 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     6781 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     6841 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     6901 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     6961 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     7021 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     7081 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     7141 tttgctcaca tggctcgaca
//
