LOCUS       sequence-466945-        6235 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 6235)
  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 6235)
  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-142
FEATURES             Location/Qualifiers
     source          1..6235
                     /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(1348..1372)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1725..1766
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(1806..1825)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1850..1898
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      2100..2528
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2187..2206)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2397..2418
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2537..2557)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2542..2871
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2722..2857
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2784..2803
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2938..3732
                     /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(2992..3011)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3602..3621
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3908..4029
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3945..3964)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3999..4018
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4078..4094)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4078..4094)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4091..4113)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4102..4118
                     /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(4126..4132,4133..4150,4151..4156))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4171..4192
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4309..4326)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4480..5068)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4560..4579)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5239..6030,6031..6099))
                     /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     5862..5881
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6100..6204)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct 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 atgggtggag 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 ggggaggtag aggaggcaag
      961 tctggcgcca tgctgagtca ccgcccacaa ggcccagggc gggccctcgg ggggccctgg
     1021 cagggttggg gggatcttag gaagccacaa ggagggctgg ggggctcttg gagcaggagt
     1081 caggaggcct gggcagcctg aagagtacac gccgacggac agacagacag tgcagtcacc
     1141 cataaagtag aaagcactac taacagcact ggagggtgta gtgtttccta ctttatggat
     1201 gagtgtactg tgggcttcgg agatcacgcc actgctgccg cccgctgccc gccaccatct
     1261 tcctcggcgc tcggggacct cgtgtgacag gtgagcacct tacggcccct ccctaccctg
     1321 cccagatgcc tgaaaggcct caagcttacc cagctttctt gtacaaagtg gttgatctag
     1381 agggcccgca gtggggtcag cttctaccgg ggcggcgtcc cggggtcttg gaactgcatg
     1441 cagactgcct gcttgggaaa catacttctt tatatgccca tatggacctg ctaagctatg
     1501 gaatgtaaag aagtatgtat ctcaggccgg gacctctctc gccgcactga ggggcactcc
     1561 acaccacggg ggccgccgcg gccgacagcc agccaggcgg ctgtggtggg ggtgaggctg
     1621 gggcggcagg ctgcctgggc gttggtggac ggtcgtgtcc ccgccagcat tgttctgtca
     1681 ctttgcctgt gtcagcaggt ggaagtgctc accagacatt cgaaggtaag cctatcccta
     1741 accctctcct cggtctcgat tctacgcgta ccggttagta atgagtttaa acgggggagg
     1801 ctaactgaaa cacggaagga gacaataccg gaaggaaccc gcgctatgac ggcaataaaa
     1861 agacagaata aaacgcacgg gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca
     1921 gggctggcac tctgtcgata ccccaccgag accccattgg ggccaatacg cccgcgtttc
     1981 ttccttttcc ccaccccacc ccccaagttc gggtgaaggc ccagggctcg cagccaacgt
     2041 cggggcggca ggccctgcca tagcagatct gcgcagctgg ggctctaggg ggtatcccca
     2101 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     2161 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     2221 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     2281 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     2341 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     2401 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     2461 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     2521 cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     2581 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     2641 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     2701 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     2761 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag
     2821 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg
     2881 ggagcttgta tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg
     2941 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     3001 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     3061 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag
     3121 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     3181 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     3241 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     3301 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     3361 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     3421 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc
     3481 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     3541 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     3601 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     3661 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     3721 gagttcttct gagcgggact ctggggttcg cgaaatgacc gaccaagcga cgcccaacct
     3781 gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt
     3841 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc
     3901 ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     3961 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     4021 tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc
     4081 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
     4141 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     4201 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     4261 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     4321 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     4381 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     4441 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     4501 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     4561 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     4621 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     4681 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     4741 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     4801 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     4861 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     4921 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     4981 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     5041 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     5101 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     5161 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     5221 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     5281 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     5341 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     5401 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     5461 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     5521 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     5581 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     5641 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     5701 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     5761 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     5821 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     5881 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     5941 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     6001 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     6061 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     6121 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     6181 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
