LOCUS       sequence-142921-        6639 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 6639)
  AUTHORS   O'Donnell KA, Wentzel EA, Zeller KI, Dang CV, Mendell JT
  TITLE     c-Myc-regulated microRNAs modulate E2F1 expression.
  JOURNAL   Nature. 2005 Jun 9. 435(7043):839-43.
  PUBMED    15944709
REFERENCE   2  (bases 1 to 6639)
  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. 
            2005 Jun 9. 435(7043):839-43."
COMMENT     Sequence Label: pcDNA3.1/V5-His-TOPO-mir17-92
FEATURES             Location/Qualifiers
     source          1..6639
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(63..80)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(234..822)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(314..333)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(993..1784,1785..1853))
                     /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     1616..1635
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1854..1958)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2033..2052)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2224..2603
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2604..2807
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2758..2778
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2852..2871
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2852..2870
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             4126..4167
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             4177..4194
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4217..4234)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4223..4447
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4493..4921
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4580..4599)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4790..4811
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4930..4950)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4935..5264
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5115..5250
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5177..5196
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5331..6125
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5385..5404)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5995..6014
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6301..6422
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6338..6357)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6392..6411
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6471..6487)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6471..6487)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6484..6506)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6495..6511
                     /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(6519..6525,6526..6543,6544..6549))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6564..6585
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
       61 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
      121 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
      181 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
      241 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
      301 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
      361 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
      421 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
      481 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
      541 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
      601 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
      661 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
      721 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
      781 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
      841 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
      901 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
      961 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     1021 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     1081 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     1141 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     1201 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     1261 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     1321 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     1381 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     1441 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     1501 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     1561 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     1621 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     1681 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     1741 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     1801 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     1861 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     1921 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     1981 cctgacgtcg acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct
     2041 gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg
     2101 agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga
     2161 agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc
     2221 gttgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
     2281 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
     2341 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
     2401 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
     2461 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
     2521 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
     2581 tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat
     2641 agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt
     2701 tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc
     2761 aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctctc tggctaacta
     2821 gagaacccac tgcttactgg cttatcgaaa ttaatacgac tcactatagg gagacccaag
     2881 ctggctagtt aagcttggta ccgagctcgg atccactagt ccagtgtggt ggaattgccc
     2941 ttccggaatt cctaaatgga cctcatatct ttgagataat taaactaatt ttttcttccc
     3001 cattagggat tatgctgaat ttgtatggtt tatagttgtt agagtttgag gtgttaattc
     3061 taattatcta tttcaaattt agcaggaaaa aagagaacat caccttgtaa aactgaagat
     3121 tgtgaccagt cagaataatg tcaaagtgct tacagtgcag gtagtgatat gtgcatctac
     3181 tgcagtgaag gcacttgtag cattatggtg acagctgcct cgggaagcca agttgggctt
     3241 taaagtgcag ggcctgctga tgttgagtgc tttttgttct aaggtgcatc tagtgcagat
     3301 agtgaagtag attagcatct actgccctaa gtgctccttc tggcataaga agttatgtat
     3361 tcatccaata attcaagcca agcaagtata taggtgtttt aatagttttt gtttgcagtc
     3421 ctctgttagt tttgcatagt tgcactacaa gaagaatgta gttgtgcaaa tctatgcaaa
     3481 actgatggtg gcctgctatt tccttcaaat gaatgatttt tactaatttt gtgtactttt
     3541 attgtgtcga tgtagaatct gcctggtcta tctgatgtga cagcttctgt agcactaaag
     3601 tgcttatagt gcaggtagtg tttagttatc tactgcatta tgagcactta aagtactgct
     3661 agctgtagaa ctccagcttc ggcctgtcgc ccaatcaaac tgtcctgtta ctgaacactg
     3721 ttctatggtt agttttgcag gtttgcatcc agctgtgtga tattctgctg tgcaaatcca
     3781 tgcaaaactg actgtggtag tgaaaagtct gtagaaaagt aagggaaact caaacccctt
     3841 tctacacagg ttgggatcgg ttgcaatgct gtgtttctgt atggtattgc acttgtcccg
     3901 gcctgttgag tttggtgggg attgtgacca gaagattttg aaaattaaat attactgaag
     3961 atttcgactt ccactgttaa atgtacaaga tacatgaaat attaaagaaa atgtgtaact
     4021 ttttgtgtaa atacatcttg tcttgttttc gaattccgga agggcaattc tgcagatatc
     4081 cagcacagtg gcggccgctc gagtctagag ggcccgcggt tcgaaggtaa gcctatccct
     4141 aaccctctcc tcggtctcga ttctacgcgt accggtcatc atcaccatca ccattgagtt
     4201 taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc
     4261 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat
     4321 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg
     4381 caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc
     4441 tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc
     4501 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     4561 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     4621 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     4681 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
     4741 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     4801 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     4861 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     4921 taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     4981 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     5041 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     5101 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     5161 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc
     5221 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt
     5281 gtatatccat tttcggatct gatcaagaga caggatgagg atcgtttcgc atgattgaac
     5341 aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact
     5401 gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc
     5461 gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg
     5521 cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg
     5581 tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt
     5641 catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc
     5701 atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag
     5761 cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg
     5821 ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc
     5881 tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt
     5941 ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg
     6001 ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt
     6061 acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct
     6121 tctgagcggg actctggggt tcgcgaaatg accgaccaag cgacgcccaa cctgccatca
     6181 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
     6241 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc
     6301 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     6361 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     6421 tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg
     6481 tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata
     6541 aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca
     6601 ctgcccgctt tccagtcggg aaacctgtcg tgccagctg
//
