LOCUS       sequence-148202-        5810 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 5810)
  AUTHORS   Chiang HR, Schoenfeld LW, Ruby JG, Auyeung VC, Spies N, Baek D, 
            Johnston WK, Russ C, Luo S, Babiarz JE, Blelloch R, Schroth GP, 
            Nusbaum C, Bartel DP
  TITLE     Mammalian microRNAs: experimental evaluation of novel and previously
            annotated genes.
  JOURNAL   Genes Dev. 2010 May 15. 24(10):992-1009.
  PUBMED    20413612
REFERENCE   2  (bases 1 to 5810)
  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"; journalName: "Genes Dev. 
            2010 May 15. 24(10):992-1009."
COMMENT     Sequence Label: pcDNA3.2/V5 hsa-mir-9-1
FEATURES             Location/Qualifiers
     source          1..5810
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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)."
     primer_bind     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    270..391
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(281..300)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     335..354
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(567..1361)
                     /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(678..697)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1288..1307
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(1428..1757)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1442..1577
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1496..1515)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     1742..1762
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(1771..2199)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1881..1902)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2093..2112
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    2401..2449
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     2474..2493
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(2533..2574)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     protein_bind    2600..2624
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2939..2963)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2992..3011)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2993..3011)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(3056..3259)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(3085..3105)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        3260..3639
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     3811..3830
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        3905..4009
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4010..4078,4079..4870)
                     /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     complement(4228..4247)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5041..5629
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5530..5549
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5783..5800
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctct
      241 agctagaggt cgacggtata cagacatgat aagatacatt gatgagtttg gacaaaccac
      301 aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt
      361 tgtaaccatt ataagctgca ataaacaagt tggggtgggc gaagaactcc agcatgagat
      421 ccccgcgctg gaggatcatc cagccggcgt cccggaaaac gattccgaag cccaaccttt
      481 catagaaggc ggcggtggaa tcgaaatctc gtgatggcag gttgggcgtc gcttggtcgg
      541 tcatttcgcg aaccccagag tcccgctcag aagaactcgt caagaaggcg atagaaggcg
      601 atgcgctgcg aatcgggagc ggcgataccg taaagcacga ggaagcggtc agcccattcg
      661 ccgccaagct cttcagcaat atcacgggta gccaacgcta tgtcctgata gcggtccgcc
      721 acacccagcc ggccacagtc gatgaatcca gaaaagcggc cattttccac catgatattc
      781 ggcaagcagg catcgccatg ggtcacgacg agatcctcgc cgtcgggcat gcgcgccttg
      841 agcctggcga acagttcggc tggcgcgagc ccctgatgct cttcgtccag atcatcctga
      901 tcgacaagac cggcttccat ccgagtacgt gctcgctcga tgcgatgttt cgcttggtgg
      961 tcgaatgggc aggtagccgg atcaagcgta tgcagccgcc gcattgcatc agccatgatg
     1021 gatactttct cggcaggagc aaggtgagat gacaggagat cctgccccgg cacttcgccc
     1081 aatagcagcc agtcccttcc cgcttcagtg acaacgtcga gcacagctgc gcaaggaacg
     1141 cccgtcgtgg ccagccacga tagccgcgct gcctcgtcct gcagttcatt cagggcaccg
     1201 gacaggtcgg tcttgacaaa aagaaccggg cgcccctgcg ctgacagccg gaacacggcg
     1261 gcatcagagc agccgattgt ctgttgtgcc cagtcatagc cgaatagcct ctccacccaa
     1321 gcggccggag aacctgcgtg caatccatct tgttcaatca tgcgaaacga tcctcatcct
     1381 gtctcttgat cagatccgaa aatggatata caagctcccg ggagcttttt gcaaaagcct
     1441 aggcctccaa aaaagcctcc tcactacttc tggaatagct cagaggcaga ggcggcctcg
     1501 gcctctgcat aaataaaaaa aattagtcag ccatggggcg gagaatgggc ggaactgggc
     1561 ggagttaggg gcgggatggg cggagttagg ggcgggacta tggttgctga ctaattgaga
     1621 tgcatgcttt gcatacttct gcctgctggg gagcctgggg actttccaca cctggttgct
     1681 gactaattga gatgcatgct ttgcatactt ctgcctgctg gggagcctgg ggactttcca
     1741 caccctaact gacacacatt ccacagaatt aattcgcgtt aaatttttgt taaatcagct
     1801 cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg
     1861 agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact
     1921 ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac
     1981 cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga
     2041 gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga
     2101 aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca
     2161 ccacacccgc cgcgcttaat gcgccgctac agggcgcgtg gggatacccc ctagagcccc
     2221 agctgcgcag atctgctatg gcagggcctg ccgccccgac gttggctgcg agccctgggc
     2281 cttcacccga acttgggggg tggggtgggg aaaaggaaga aacgcgggcg tattggcccc
     2341 aatggggtct cggtggggta tcgacagagt gccagccctg ggaccgaacc ccgcgtttat
     2401 gaacaaacga cccaacaccc gtgcgtttta ttctgtcttt ttattgccgt catagcgcgg
     2461 gttccttccg gtattgtctc cttccgtgtt tcagttagcc tcccccgttt aaactcatta
     2521 ctaaccggta cgcgtagaat cgagaccgag gagagggtta gggataggct taccttcgaa
     2581 ccgcgggccc tctagatcaa ccactttgta caagaaagct gggtaagctt atcctctggt
     2641 gctggtcagt cctcgcggct ctagcccctc gctcaagcct ctccctcccg cccgtgaggc
     2701 cggctttccc cgggcccctc tgcgcagtgt atggggttat ttttactttc ggttatctag
     2761 ctttatgaag actccacacc actcatacag ctagataacc aaagataaca accaaccccg
     2821 cctcctggct gctgtcgccg cctcttccac gcagcctccc ggccgccgcc gccgccagca
     2881 cctccgcagc ttcccggtcg cccgtcagcg ggagtaggag ggaagggaca cgcatatgag
     2941 cctgcttttt tgtacaaact tgttgatagc ttaactagcc agcttgggtc tccctatagt
     3001 gagtcgtatt aatttcgata agccagtaag cagtgggttc tctagttagc cagagagctc
     3061 tgcttatata gacctcccac cgtacacgcc taccgcccat ttgcgtcaat ggggcggagt
     3121 tgttacgaca ttttggaaag tcccgttgat tttggtgcca aaacaaactc ccattgacgt
     3181 caatggggtg gagacttgga aatccccgtg agtcaaaccg ctatccacgc ccattgatgt
     3241 actgccaaaa ccgcatcacc atggtaatag cgatgactaa tacgtagatg tactgccaag
     3301 taggaaagtc ccataaggtc atgtactggg cataatgcca ggcgggccat ttaccgtcat
     3361 tgacgtcaat agggggcgta cttggcatat gatacacttg atgtactgcc aagtgggcag
     3421 tttaccgtaa atactccacc cattgacgtc aatggaaagt ccctattggc gttactatgg
     3481 gaacatacgt cattattgac gtcaatgggc gggggtcgtt gggcggtcag ccaggcgggc
     3541 catttaccgt aagttatgta acgcggaact ccatatatgg gctatgaact aatgaccccg
     3601 taattgatta ctattaataa ctagtcaata atcaatgtca acgcgtatat ctggcccgta
     3661 catcgcgaag cagcgcaaaa cgcctaaccc taagcagatt cttcatgcaa ttgtcggtca
     3721 agccttgcct tgttgtagct taaattttgc tcgcgcacta ctcagcgacc tccaacacac
     3781 aagcagggag cagatactgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
     3841 accatagggg atcgggagat ctcccgatcc gtcgacgtca ggtggcactt ttcggggaaa
     3901 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     3961 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     4021 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     4081 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     4141 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     4201 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     4261 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     4321 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     4381 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     4441 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     4501 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     4561 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     4621 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     4681 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     4741 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     4801 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     4861 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     4921 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     4981 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     5041 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     5101 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     5161 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     5221 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     5281 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     5341 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     5401 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     5461 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     5521 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     5581 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     5641 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     5701 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     5761 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
//
