LOCUS       sequence-117478-        6006 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 6006)
  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 6006)
  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 mir50-wt
FEATURES             Location/Qualifiers
     source          1..6006
                     /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(1119..1143)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1496..1537
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(1577..1596)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1621..1669
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      1871..2299
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1958..1977)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2168..2189
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2308..2328)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2313..2642
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2493..2628
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2555..2574
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2709..3503
                     /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(2763..2782)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3373..3392
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3679..3800
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3716..3735)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3770..3789
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3849..3865)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3849..3865)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3862..3884)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3873..3889
                     /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(3897..3903,3904..3921,3922..3927))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3942..3963
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4080..4097)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4251..4839)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4331..4350)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5010..5801,5802..5870))
                     /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     5633..5652
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5871..5975)
                     /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 aggctctatt tttactaatt ctgaattcca
      961 taattctata ttctgtattc ctgcccgccg gccgctgata tgtctggtat tcttgggttt
     1021 gaacttccag cgttgaaccc gcatattaga cgtatcgacg gccggcgggg caggtaatgg
     1081 ctggtaagtt tggagagacc gggggagtcc cagagatgcc ccagctttct tgtacaaagt
     1141 ggttgatcta gagggcccgc agtggggtca gcttctaccg gggcggcgtc ccggggtctt
     1201 ggaactgcat gcagactgcc tgcttgggaa acatacttct ttatatgccc atatggacct
     1261 gctaagctat ggaatgtaaa gaagtatgta tctcaggccg ggacctctct cgccgcactg
     1321 aggggcactc cacaccacgg gggccgccgc ggccgacagc cagccaggcg gctgtggtgg
     1381 gggtgaggct ggggcggcag gctgcctggg cgttggtgga cggtcgtgtc cccgccagca
     1441 ttgttctgtc actttgcctg tgtcagcagg tggaagtgct caccagacat tcgaaggtaa
     1501 gcctatccct aaccctctcc tcggtctcga ttctacgcgt accggttagt aatgagttta
     1561 aacgggggag gctaactgaa acacggaagg agacaatacc ggaaggaacc cgcgctatga
     1621 cggcaataaa aagacagaat aaaacgcacg ggtgttgggt cgtttgttca taaacgcggg
     1681 gttcggtccc agggctggca ctctgtcgat accccaccga gaccccattg gggccaatac
     1741 gcccgcgttt cttccttttc cccaccccac cccccaagtt cgggtgaagg cccagggctc
     1801 gcagccaacg tcggggcggc aggccctgcc atagcagatc tgcgcagctg gggctctagg
     1861 gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     1921 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     1981 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcggggcat ccctttaggg
     2041 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     2101 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     2161 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     2221 tttgatttat aagggatttt ggggatttcg gcctattggt taaaaaatga gctgatttaa
     2281 caaaaattta acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc
     2341 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt
     2401 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     2461 cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg
     2521 cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct
     2581 ctgcctctga gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca
     2641 aaaagctccc gggagcttgt atatccattt tcggatctga tcaagagaca ggatgaggat
     2701 cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga
     2761 ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc
     2821 ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga
     2881 atgaactgca ggacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg
     2941 cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc
     3001 cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg
     3061 atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga
     3121 aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc
     3181 tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca
     3241 tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg
     3301 tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct
     3361 atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg
     3421 accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc
     3481 gccttcttga cgagttcttc tgagcgggac tctggggttc gcgaaatgac cgaccaagcg
     3541 acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     3601 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     3661 gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     3721 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     3781 aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg
     3841 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     3901 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     3961 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     4021 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     4081 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     4141 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     4201 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4261 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4321 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     4381 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     4441 tctcaatgct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     4501 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     4561 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     4621 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     4681 tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     4741 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     4801 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     4861 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     4921 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     4981 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     5041 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     5101 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     5161 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     5221 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     5281 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     5341 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     5401 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     5461 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     5521 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     5581 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     5641 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     5701 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     5761 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     5821 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     5881 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     5941 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     6001 gacgtc
//
