LOCUS       sequence-66030-e        6218 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 6218)
  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 6218)
  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 dme-mir-92a
FEATURES             Location/Qualifiers
     source          1..6218
                     /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(1331..1355)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1708..1749
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(1789..1808)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1833..1881
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      2083..2511
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2170..2189)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2380..2401
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2520..2540)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2525..2854
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2705..2840
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2767..2786
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2921..3715
                     /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(2975..2994)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3585..3604
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3891..4012
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3928..3947)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3982..4001
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4061..4077)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4061..4077)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4074..4096)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4085..4101
                     /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(4109..4115,4116..4133,4134..4139))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4154..4175
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4292..4309)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4463..5051)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4543..4562)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5222..6013,6014..6082))
                     /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     5845..5864
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6083..6187)
                     /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 aggctttgaa attcgagatt tttgggcttt
      961 ggtcgatgct cctattgttc gcctacctgc ctacctgttg atggtattat ttgtgaatgt
     1021 ttatcaagat gttccatgta cttgctgaaa ctcaatattc gtgatctgat tttcagccga
     1081 atataaatat gaatttcccg taggacggga aggtgtcaac gttttgcatt tcgaataaac
     1141 attgcacttg tcccggccta tgggcggttt gtaataaaca actaaaatct ataaaattta
     1201 tttattatta ctaaagtcat gtctcgtatt tttgcttatg aaatattttc tgttgtttca
     1261 gatctcgaat tcgcaatatc agaacatctg tgttatttta attatataaa agtatatata
     1321 caggtgagtt acccagcttt cttgtacaaa gtggttgatc tagagggccc gcagtggggt
     1381 cagcttctac cggggcggcg tcccggggtc ttggaactgc atgcagactg cctgcttggg
     1441 aaacatactt ctttatatgc ccatatggac ctgctaagct atggaatgta aagaagtatg
     1501 tatctcaggc cgggacctct ctcgccgcac tgaggggcac tccacaccac gggggccgcc
     1561 gcggccgaca gccagccagg cggctgtggt gggggtgagg ctggggcggc aggctgcctg
     1621 ggcgttggtg gacggtcgtg tccccgccag cattgttctg tcactttgcc tgtgtcagca
     1681 ggtggaagtg ctcaccagac attcgaaggt aagcctatcc ctaaccctct cctcggtctc
     1741 gattctacgc gtaccggtta gtaatgagtt taaacggggg aggctaactg aaacacggaa
     1801 ggagacaata ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca
     1861 cgggtgttgg gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg cactctgtcg
     1921 ataccccacc gagaccccat tggggccaat acgcccgcgt ttcttccttt tccccacccc
     1981 accccccaag ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg
     2041 ccatagcaga tctgcgcagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg
     2101 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc
     2161 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     2221 gtcaagctct aaatcggggc atccctttag ggttccgatt tagtgcttta cggcacctcg
     2281 accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     2341 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     2401 gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttggggattt
     2461 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg
     2521 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     2581 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
     2641 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     2701 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     2761 tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg
     2821 aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat
     2881 tttcggatct gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt
     2941 gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca
     3001 gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct
     3061 ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct
     3121 atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc
     3181 gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct
     3241 tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga
     3301 tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg
     3361 gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc
     3421 agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac
     3481 ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat
     3541 cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga
     3601 tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc
     3661 cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg
     3721 actctggggt tcgcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg
     3781 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     3841 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta
     3901 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     3961 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     4021 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     4081 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     4141 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     4201 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     4261 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     4321 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     4381 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     4441 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     4501 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     4561 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     4621 ccgcctttct cccttcggga agcgtggcgc tttctcaatg ctcacgctgt aggtatctca
     4681 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     4741 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     4801 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     4861 cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct
     4921 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     4981 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     5041 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     5101 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     5161 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     5221 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     5281 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     5341 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     5401 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     5461 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     5521 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     5581 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     5641 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     5701 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     5761 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     5821 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     5881 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     5941 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     6001 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     6061 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     6121 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     6181 ttccgcgcac atttccccga aaagtgccac ctgacgtc
//
