LOCUS       sequence-117350-        6013 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 6013)
  AUTHORS   Auyeung VC, Ulitsky I, McGeary SE, Bartel DP
  TITLE     Beyond Secondary Structure: Primary-Sequence Determinants License 
            Pri-miRNA Hairpins for Processing.
  JOURNAL   Cell. 2013 Feb 14;152(4):844-58. doi: 10.1016/j.cell.2013.01.031.
  PUBMED    23415231
REFERENCE   2  (bases 1 to 6013)
  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: 
            "10.1016/j.cell.2013.01"; journalName: "Cell"; date: "2013-02-14- 
            14"; volume: "152"; issue: "4"; pages: "844-58"
COMMENT     Sequence Label: pcDNA3.2 mir-1-1 reporter mir28-1
FEATURES             Location/Qualifiers
     source          1..6013
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    911..935
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(1126..1150)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1503..1544
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(1584..1603)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1628..1676
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      1878..2306
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1965..1984)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2175..2196
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2315..2335)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2320..2649
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2500..2635
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2562..2581
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2716..3510
                     /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(2770..2789)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3380..3399
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3686..3807
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3723..3742)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3777..3796
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3856..3872)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3856..3872)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3869..3891)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3880..3896
                     /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(3904..3910,3911..3928,3929..3934))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3949..3970
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4087..4104)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4258..4846)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4338..4357)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5017..5808,5809..5877))
                     /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     5640..5659
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5878..5982)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagt
      901 taagctatca acaagtttgt acaaaaaagc aggctggtag acaacatgct tgatgctgaa
      961 gatacaagaa aaaatttaaa acagtccttg ccctcaagga gctcacagtc tattgagtta
     1021 cctttctgac tttcccacta gattgtgagc tcctggaggg caggcacttt cgttcatctg
     1081 aaaaagagct taaatttcag tgttaatcct agattacaat cccgccccca gctttcttgt
     1141 acaaagtggt tgatctagag ggcccgcagt ggggtcagct tctaccgggg cggcgtcccg
     1201 gggtcttgga actgcatgca gactgcctgc ttgggaaaca tacttcttta tatgcccata
     1261 tggacctgct aagctatgga atgtaaagaa gtatgtatct caggccggga cctctctcgc
     1321 cgcactgagg ggcactccac accacggggg ccgccgcggc cgacagccag ccaggcggct
     1381 gtggtggggg tgaggctggg gcggcaggct gcctgggcgt tggtggacgg tcgtgtcccc
     1441 gccagcattg ttctgtcact ttgcctgtgt cagcaggtgg aagtgctcac cagacattcg
     1501 aaggtaagcc tatccctaac cctctcctcg gtctcgattc tacgcgtacc ggttagtaat
     1561 gagtttaaac gggggaggct aactgaaaca cggaaggaga caataccgga aggaacccgc
     1621 gctatgacgg caataaaaag acagaataaa acgcacgggt gttgggtcgt ttgttcataa
     1681 acgcggggtt cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg
     1741 ccaatacgcc cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc
     1801 agggctcgca gccaacgtcg gggcggcagg ccctgccata gcagatctgc gcagctgggg
     1861 ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     1921 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
     1981 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc ggggcatccc
     2041 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
     2101 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     2161 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
     2221 ctattctttt gatttataag ggattttggg gatttcggcc tattggttaa aaaatgagct
     2281 gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga
     2341 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     2401 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     2461 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     2521 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     2581 gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     2641 ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca agagacagga
     2701 tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg
     2761 gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc
     2821 gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt
     2881 gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt
     2941 ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc
     3001 gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc
     3061 atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac
     3121 caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag
     3181 gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag
     3241 gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat
     3301 atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg
     3361 gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa
     3421 tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc
     3481 ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgcg aaatgaccga
     3541 ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag
     3601 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     3661 catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata
     3721 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     3781 tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag
     3841 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     3901 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     3961 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     4021 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     4081 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     4141 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     4201 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     4261 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     4321 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     4381 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     4441 ggcgctttct caatgctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     4501 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     4561 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     4621 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     4681 ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     4741 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     4801 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     4861 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     4921 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     4981 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     5041 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     5101 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     5161 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     5221 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     5281 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     5341 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     5401 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     5461 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     5521 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     5581 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     5641 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     5701 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     5761 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     5821 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     5881 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     5941 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     6001 gccacctgac gtc
//
