LOCUS       sequence-117479-        6005 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 6005)
  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 6005)
  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 mir129-2-2
FEATURES             Location/Qualifiers
     source          1..6005
                     /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(1118..1142)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1495..1536
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(1576..1595)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1620..1668
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      1870..2298
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1957..1976)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2167..2188
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2307..2327)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2312..2641
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2492..2627
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2554..2573
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2708..3502
                     /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(2762..2781)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3372..3391
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3678..3799
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3715..3734)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3769..3788
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3848..3864)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3848..3864)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3861..3883)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3872..3888
                     /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(3896..3902,3903..3920,3921..3926))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3941..3962
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4079..4096)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4250..4838)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4330..4349)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5009..5800,5801..5869))
                     /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     5632..5651
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5870..5974)
                     /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 aggctcatcc tgggctgaag gcggcggcga
      961 accgaagaag ccggcatatt ctgcccttcg cgaatctttt tgcggtctgg gcttgctgta
     1021 cataactcaa tagccggaag cccttacccc aaaaagcatt tgcggagggc gcactagtag
     1081 agaagacggc agccatccag cgatcgccga agcccgcacc cagctttctt gtacaaagtg
     1141 gttgatctag agggcccgca gtggggtcag cttctaccgg ggcggcgtcc cggggtcttg
     1201 gaactgcatg cagactgcct gcttgggaaa catacttctt tatatgccca tatggacctg
     1261 ctaagctatg gaatgtaaag aagtatgtat ctcaggccgg gacctctctc gccgcactga
     1321 ggggcactcc acaccacggg ggccgccgcg gccgacagcc agccaggcgg ctgtggtggg
     1381 ggtgaggctg gggcggcagg ctgcctgggc gttggtggac ggtcgtgtcc ccgccagcat
     1441 tgttctgtca ctttgcctgt gtcagcaggt ggaagtgctc accagacatt cgaaggtaag
     1501 cctatcccta accctctcct cggtctcgat tctacgcgta ccggttagta atgagtttaa
     1561 acgggggagg ctaactgaaa cacggaagga gacaataccg gaaggaaccc gcgctatgac
     1621 ggcaataaaa agacagaata aaacgcacgg gtgttgggtc gtttgttcat aaacgcgggg
     1681 ttcggtccca gggctggcac tctgtcgata ccccaccgag accccattgg ggccaatacg
     1741 cccgcgtttc ttccttttcc ccaccccacc ccccaagttc gggtgaaggc ccagggctcg
     1801 cagccaacgt cggggcggca ggccctgcca tagcagatct gcgcagctgg ggctctaggg
     1861 ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     1921 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     1981 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcggggcatc cctttagggt
     2041 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     2101 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     2161 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt
     2221 ttgatttata agggattttg gggatttcgg cctattggtt aaaaaatgag ctgatttaac
     2281 aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     2341 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     2401 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     2461 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     2521 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     2581 tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     2641 aaagctcccg ggagcttgta tatccatttt cggatctgat caagagacag gatgaggatc
     2701 gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag
     2761 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg
     2821 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa
     2881 tgaactgcag gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc
     2941 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc
     3001 ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga
     3061 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa
     3121 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct
     3181 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat
     3241 gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt
     3301 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta
     3361 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga
     3421 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg
     3481 ccttcttgac gagttcttct gagcgggact ctggggttcg cgaaatgacc gaccaagcga
     3541 cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     3601 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     3661 agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     3721 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     3781 aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt
     3841 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     3901 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     3961 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     4021 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     4081 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     4141 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     4201 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     4261 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     4321 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     4381 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     4441 ctcaatgctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     4501 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     4561 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     4621 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     4681 acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     4741 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     4801 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     4861 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     4921 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     4981 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     5041 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     5101 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     5161 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     5221 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     5281 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     5341 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     5401 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     5461 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     5521 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     5581 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     5641 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     5701 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     5761 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     5821 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     5881 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     5941 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     6001 acgtc
//
