LOCUS       sequence-12517-e        7031 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 7031)
  AUTHORS   Trujillo RD, Yue SB, Tang Y, O'Gorman WE, Chen CZ
  TITLE     The potential functions of primary microRNAs in target recognition 
            and repression.
  JOURNAL   EMBO J. 2010 Oct 6;29(19):3272-85. doi: 10.1038/emboj.2010.208. Epub
            2010 Aug 31.
  PUBMED    20808284
REFERENCE   2  (bases 1 to 7031)
  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.2010.208"; journalName: "EMBO J"; date: "2010-10-6- 
            6"; volume: "29"; issue: "19"; pages: "3272-85"
COMMENT     Sequence Label: MDH1-pre-let-7hairpin-PGK-GFP
FEATURES             Location/Qualifiers
     source          1..7031
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    580..921
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             988..1404
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1334..1356
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1372..1388
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     primer_bind     1716..1732
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     promoter        1796..2296
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     2229..2248
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             join(2306..2308,2309..2311,2312..3025)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2351..2372)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2612..2631)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2959..2980
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(3236..3326)
                     /label=H1 promoter
                     /note="human H1 RNA promoter"
     primer_bind     complement(3296..3315)
                     /label=H1
                     /note="Human H1 promoter, forward primer"
     primer_bind     complement(3990..4006)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3990..4006)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4003..4025)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4014..4030
                     /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(4038..4044,4045..4062,4063..4068))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4083..4104
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4221..4238)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4392..4980)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4472..4491)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5151..5942,5943..6011))
                     /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     5774..5793
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6012..6116)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6184..6202
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6240..6262)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6362..6381
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6575..6597
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6589..6606
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6590..6606
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6809..6828
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             join(7030..7031,1..516)
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
ORIGIN
        1 tgaaagaccc cacctgtagg tttggcaagc tagcttaagt aacgccattt tgcaaggcat
       61 ggaaaataca taactgagaa tagagaagtt cagatcaagg ttaggaacag agagacagca
      121 gaatatgggc caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga
      181 acagatggtc cccagatgcg gtcccgccct cagcagtttc tagagaacca tcagatgttt
      241 ccagggtgcc ccaaggacct gaaaatgacc ctgtgcctta tttgaactaa ccaatcagtt
      301 cgcttctcgc ttctgttcgc gcgcttctgc tccccgagct caataaaaga gcccacaacc
      361 cctcactcgg cgcgccagtc ctccgataga ctgcgtcgcc cgggtacccg tattcccaat
      421 aaagcctctt gctgtttgca tccgaatcgt ggactcgctg atccttggga gggtctcctc
      481 agattgattg actgcccacc tcgggggtct ttcatttgga ggttccaccg agatttggag
      541 acccctgcct agggaccacc gacccccccg ccgggaggta agctggccag cggtcgtttc
      601 gtgtctgtct ctgtctttgt gcgtgtttgt gccggcatct aatgtttgcg cctgcgtctg
      661 tactagttag ctaactagct ctgtatctgg cggacccgtg gtggaactga cgagttctga
      721 acacccggcc gcaaccctgg gagacgtccc agggactttg ggggccgttt ttgtggcccg
      781 acctgaggaa gggagtcgat gtggaatccg accccgtcag gatatgtggt tctggtagga
      841 gacgagaacc taaaacagtt cccgcctccg tctgaatttt tgctttcggt ttggaaccga
      901 agccgcgcgt cttgtctgct gcagcgctgc agcatcgttc tgtgttgtct ctgtctgact
      961 gtgtttctgt atttgtctga aaattagggc cagactgtta ccactccctt aagtttgacc
     1021 ttaggtcact ggaaagatgt cgagcggatc gctcacaacc agtcggtaga tgtcaagaag
     1081 agacgttggg ttaccttctg ctctgcagaa tggccaacct ttaacgtcgg atggccgcga
     1141 gacggcacct ttaaccgaga cctcatcacc caggttaaga tcaaggtctt ttcacctggc
     1201 ccgcatggac acccagacca ggtcccctac atcgtgacct gggaagcctt ggcttttgac
     1261 ccccctccct gggtcaagcc ctttgtacac cctaagcctc cgcctcctct tcctccatcc
     1321 gccccgtctc tcccccttga acctcctcgt tcgaccccgc ctcgatcctc cctttatcca
     1381 gccctcactc cttctctagg cgccggaatt agatcttata tggggcaccc ccgccccttg
     1441 taaacttccc tgaccctgac atgacaagag ttactaacag cccctctctc caagctcact
     1501 tacaggctct ctacttagtc cagcacgaag tctggagacc tctggcggca gcctaccaag
     1561 aacaactgga ccgaccggtg gtacctcacc cttaccgagt cggcgacaca gtgtgggtcc
     1621 gccgacacca gactaagaac ctagaacctc gctggaaagg accttacaca gtcctgctga
     1681 ccacccccac cgccctcaaa gtagacggca tcgcagcttg gatacacgcc gcccacgtga
     1741 aggctgccga ccccgggggt ggaccatcct ctagactgcc ggatcaattc ctaccgggta
     1801 ggggaggcgc ttttcccaag gcagtctgga gcatgcgctt tagcggcccc gctgggcact
     1861 tggcgctaca caagtggcct ctggcctcgc acacattcca catccaccgg taggcgccaa
     1921 ccggctccgt tctttggtgg ccccttcgcg ccaccttcta ctcctcccct agtcaggaag
     1981 ttcccccccg ccccgcagct cgcgtcgtgc aggacgtgac aaatggaagt agcacgtctc
     2041 actagtctcg tgcagatgga cagcaccgct gagcaatgga agcgggtagg cctttggggc
     2101 agcggccaat agcagctttg gctccttcgc tttctgggct cagaggctgg gaaggggtgg
     2161 gtccgggggc gggctcaggg gcgggctcag gggcggggcg ggcgcccgaa ggtcctccgg
     2221 aggcccggca ttctgcacgc ttcaaaagcg cacgtctgcc gcgctgttct cctcttcctc
     2281 atctccgggc ctttcgacct gcaccatggt gagcaagggc gaggagctgt tcaccggggt
     2341 ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg
     2401 cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg
     2461 caagctgccc gtgccctggc ccaccctcgt gaccaccttc acctacggcg tgcagtgctt
     2521 cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg
     2581 ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga
     2641 ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa
     2701 ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta
     2761 tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat
     2821 cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg
     2881 ccccgtgctg ctgcccgaca accactacct gagcacccag tccgccctga gcaaagaccc
     2941 caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct
     3001 cggcatggac gagctgtaca agtaatgaat taattaagaa ttgcggccgc gtcgacctgc
     3061 agccaagctt atcgataaaa taaaagattt tatttagtct ccagaaaaag gggggaatga
     3121 aagaccccac ctgtaggttt ggcaagaatt cttaattaaa aaggtaaggt agaaaattgc
     3181 atagttcacc ggtggtaata ttccaaacta tacaacctac tacctcaggg ctcgagtggt
     3241 ctcatacaga acttataaga ttcccaaatc caaagacatt tcacgtttat ggtgatttcc
     3301 cagaacacat agcgacatgc aaatattggg atccgctagc ttaagtaacg ccattttgca
     3361 aggcatggaa aatacataac tgagaataga gaagttcaga tcaaggttag gaacagagag
     3421 acagcagaat atgggccaaa caggatatct gtggtaagca gttcctgccc cggctcaggg
     3481 ccaagaacag atggtcccca gatgcggtcc cgccctcagc agtttctaga gaaccatcag
     3541 atgtttccag ggtgccccaa ggacctgaaa atgaccctgt gccttatttg aactaaccaa
     3601 tcagttcgct tctcgcttct gttcgcgcgc ttctgctccc cgagctcaat aaaagagccc
     3661 acaacccctc actcggcgcg ccagtcctcc gatagactgc gtcgcccggg tacccgtgta
     3721 tccaataaac cctcttgcag ttgcatccga cttgtggtct cgctgttcct tgggagggtc
     3781 tcctctgagt gattgactac ccgtcagcgg gggtctttca tgggtaacag tttcttgaag
     3841 ttggagaaca acattctgag ggtaggagtc gaatattaag taatcctgac tcaattagcc
     3901 actgttttga atccacatac tccaatactc ctgaaatagt tcattatgga cagcgcagaa
     3961 gagctgggga gaattaattc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     4021 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     4081 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     4141 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     4201 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     4261 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     4321 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     4381 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     4441 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     4501 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     4561 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     4621 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     4681 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     4741 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     4801 aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg cgctctgctg
     4861 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     4921 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     4981 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     5041 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     5101 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     5161 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     5221 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     5281 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     5341 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     5401 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     5461 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     5521 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     5581 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     5641 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     5701 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     5761 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     5821 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     5881 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     5941 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     6001 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     6061 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     6121 tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac attaacctat
     6181 aaaaataggc gtatcacgag gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac
     6241 ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc
     6301 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg gcttaactat
     6361 gcggcatcag agcagattgt actgagagtg caccatatgc ggtgtgaaat accgcacaga
     6421 tgcgtaagga gaaaataccg catcaggcgc cattcgccat tcaggctgcg caactgttgg
     6481 gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct
     6541 gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg taaaacgacg
     6601 gccagtgcca agctcgctct cccttatgcg actcctgcat taggaagcag cccagtagta
     6661 ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc atgcaaggag atggcgccca
     6721 acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg ctcatgagcc
     6781 cgaagtggcg agcccgatct tccccatcgg tgatgtcggc gatataggcg ccagcaaccg
     6841 cacctgtggc gccggtgatg ccggccacga tgcgtccggc gtagaggatc gattagtcca
     6901 atttgttaaa gacaggatat cagtggtcca ggctctagtt ttgactcaac aatatcacca
     6961 gctgaagcct atagagtacg agccatagat aaaataaaag attttattta gtctccagaa
     7021 aaagggggga a
//
