LOCUS       sequence-117375-        6226 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 6226)
  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 6226)
  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 hsa-mir-122
FEATURES             Location/Qualifiers
     source          1..6226
                     /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"
     CDS             complement(1183..1194)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     protein_bind    complement(1339..1363)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1716..1757
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(1797..1816)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1841..1889
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      2091..2519
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2178..2197)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2388..2409
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2528..2548)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2533..2862
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2713..2848
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2775..2794
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2929..3723
                     /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(2983..3002)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3593..3612
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3899..4020
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3936..3955)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3990..4009
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4069..4085)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4069..4085)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4082..4104)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4093..4109
                     /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(4117..4123,4124..4141,4142..4147))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4162..4183
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4300..4317)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4471..5059)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4551..4570)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5230..6021,6022..6090))
                     /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     5853..5872
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6091..6195)
                     /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 aggctcatat ggttcagctc ttcccattgc
      961 tcaagatgct tgtacccgtg atgcttcttt tctctgctta ggtcacaata tgtggagctg
     1021 acaaggttcc cctattatca gtgacaatgg tggaatgtgg aggtgaagtt aacaccttcg
     1081 tggctacaga gtttccttag cagagctgtg gagtgtgaca atggtgtttg tgtctaaact
     1141 atcaaacgcc attatcacac taaatagcta ctgctaggca atccttccct cgataaatgt
     1201 cttggcatcg tttgctttga gcaagaaggt tcatctgata tcagtcttct caatcttgtg
     1261 tacttactga ataaagtctg gttcttttgc actcataacc aatatggtta gctgaaccct
     1321 gcctctcaca tgaagcttac ccagctttct tgtacaaagt ggttgatcta gagggcccgc
     1381 agtggggtca gcttctaccg gggcggcgtc ccggggtctt ggaactgcat gcagactgcc
     1441 tgcttgggaa acatacttct ttatatgccc atatggacct gctaagctat ggaatgtaaa
     1501 gaagtatgta tctcaggccg ggacctctct cgccgcactg aggggcactc cacaccacgg
     1561 gggccgccgc ggccgacagc cagccaggcg gctgtggtgg gggtgaggct ggggcggcag
     1621 gctgcctggg cgttggtgga cggtcgtgtc cccgccagca ttgttctgtc actttgcctg
     1681 tgtcagcagg tggaagtgct caccagacat tcgaaggtaa gcctatccct aaccctctcc
     1741 tcggtctcga ttctacgcgt accggttagt aatgagttta aacgggggag gctaactgaa
     1801 acacggaagg agacaatacc ggaaggaacc cgcgctatga cggcaataaa aagacagaat
     1861 aaaacgcacg ggtgttgggt cgtttgttca taaacgcggg gttcggtccc agggctggca
     1921 ctctgtcgat accccaccga gaccccattg gggccaatac gcccgcgttt cttccttttc
     1981 cccaccccac cccccaagtt cgggtgaagg cccagggctc gcagccaacg tcggggcggc
     2041 aggccctgcc atagcagatc tgcgcagctg gggctctagg gggtatcccc acgcgccctg
     2101 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     2161 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     2221 ctttccccgt caagctctaa atcggggcat ccctttaggg ttccgattta gtgctttacg
     2281 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
     2341 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     2401 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
     2461 ggggatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta
     2521 attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     2581 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     2641 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     2701 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     2761 tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag
     2821 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt
     2881 atatccattt tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa
     2941 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
     3001 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
     3061 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca
     3121 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
     3181 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
     3241 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
     3301 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
     3361 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
     3421 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc
     3481 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
     3541 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
     3601 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
     3661 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
     3721 tgagcgggac tctggggttc gcgaaatgac cgaccaagcg acgcccaacc tgccatcacg
     3781 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga
     3841 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa
     3901 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     3961 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     4021 tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt
     4081 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
     4141 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     4201 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     4261 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     4321 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     4381 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     4441 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     4501 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     4561 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     4621 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcaatgct cacgctgtag
     4681 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     4741 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     4801 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     4861 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt
     4921 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     4981 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     5041 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     5101 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     5161 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     5221 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     5281 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     5341 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     5401 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     5461 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     5521 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     5581 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     5641 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     5701 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     5761 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     5821 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     5881 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     5941 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     6001 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     6061 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     6121 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     6181 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtc
//
