LOCUS       sequence-66010-e        8038 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expression of query pri-miRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8038)
  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 8038)
  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 V5-DEST mir-1-1 reporter
FEATURES             Location/Qualifiers
     source          1..8038
                     /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..1035
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(1464..1769)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(1566..1585)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(2111..2770)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2685..2704
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(2771..2873)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     protein_bind    complement(3051..3175)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             3528..3569
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(3609..3628)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3653..3701
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      3903..4331
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3990..4009)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4200..4221
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4340..4360)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4345..4674
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4525..4660
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4587..4606
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4741..5535
                     /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(4795..4814)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5405..5424
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5711..5832
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5748..5767)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5802..5821
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5881..5897)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5881..5897)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5894..5916)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5905..5921
                     /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(5929..5935,5936..5953,5954..5959))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5974..5995
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6112..6129)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6283..6871)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6363..6382)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7042..7833,7834..7902))
                     /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     7665..7684
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7903..8007)
                     /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 tgaacgagaa acgtaaaatg atataaatat
      961 caatatatta aattagattt tgcataaaaa acagactaca taatactgta aaacacaaca
     1021 tatccagtca ctatgaatca actacttaga tggtattagt gacctgtagt cgaccgacag
     1081 ccttccaaat gttcttcggg tgatgctgcc aacttagtcg accgacagcc ttccaaatgt
     1141 tcttctcaaa cggaatcgtc gtatccagcc tactcgctat tgtcctcaat gccgtattaa
     1201 atcataaaaa gaaataagaa aaagaggtgc gagcctcttt tttgtgtgac aaaataaaaa
     1261 catctaccta ttcatatacg ctagtgtcat agtcctgaaa atcatctgca tcaagaacaa
     1321 tttcacaact cttatacttt tctcttacaa gtcgttcggc ttcatctgga ttttcagcct
     1381 ctatacttac taaacgtgat aaagtttctg taatttctac tgtatcgacc tgcagactgg
     1441 ctgtgtataa gggagcctga catttatatt ccccagaaca tcaggttaat ggcgtttttg
     1501 atgtcatttt cgcggtggct gagatcagcc acttcttccc cgataacgga gaccggcaca
     1561 ctggccatat cggtggtcat catgcgccag ctttcatccc cgatatgcac caccgggtaa
     1621 agttcacggg agactttatc tgacagcaga cgtgcactgg ccagggggat caccatccgt
     1681 cgcccgggcg tgtcaataat atcactctgt acatccacaa acagacgata acggctctct
     1741 cttttatagg tgtaaacctt aaactgcatt tcaccagtcc ctgttctcgt cagcaaaaga
     1801 gccgttcatt tcaataaacc gggcgacctc agccatccct tcctgatttt ccgctttcca
     1861 gcgttcggca cgcagacgac gggcttcatt ctgcatggtt gtgcttacca gaccggagat
     1921 attgacatca tatatgcctt gagcaactga tagctgtcgc tgtcaactgt cactgtaata
     1981 cgctgcttca tagcacacct ctttttgaca tacttcgggt atacatatca gtatatattc
     2041 ttataccgca aaaatcagcg cgcaaatacg catactgtta tctggctttt agtaagccgg
     2101 atccacgcga ttacgccccg ccctgccact catcgcagta ctgttgtaat tcattaagca
     2161 ttctgccgac atggaagcca tcacagacgg catgatgaac ctgaatcgcc agcggcatca
     2221 gcaccttgtc gccttgcgta taatatttgc ccatggtgaa aacgggggcg aagaagttgt
     2281 ccatattggc cacgtttaaa tcaaaactgg tgaaactcac ccagggattg gctgagacga
     2341 aaaacatatt ctcaataaac cctttaggga aataggccag gttttcaccg taacacgcca
     2401 catcttgcga atatatgtgt agaaactgcc ggaaatcgtc gtggtattca ctccagagcg
     2461 atgaaaacgt ttcagtttgc tcatggaaaa cggtgtaaca agggtgaaca ctatcccata
     2521 tcaccagctc accgtctttc attgccatac ggaattccgg atgagcattc atcaggcggg
     2581 caagaatgtg aataaaggcc ggataaaact tgtgcttatt tttctttacg gtctttaaaa
     2641 aggccgtaat atccagctga acggtctggt tataggtaca ttgagcaact gactgaaatg
     2701 cctcaaaatg ttctttacga tgccattggg atatatcaac ggtggtatat ccagtgattt
     2761 ttttctccat tttagcttcc ttagctcctg aaaatctcga taactcaaaa aatacgcccg
     2821 gtagtgatct tatttcatta tggtgaaagt tggaacctct tacgtgccga tcaacgtctc
     2881 attttcgcca aaagttggcc cagggcttcc cggtatcaac agggacacca ggatttattt
     2941 attctgcgaa gtgatcttcc gtcacaggta tttattcggc gcaaagtgcg tcgggtgatg
     3001 ctgccaactt agtcgactac aggtcactaa taccatctaa gtagttgatt catagtgact
     3061 ggatatgttg tgttttacag tattatgtag tctgtttttt atgcaaaatc taatttaata
     3121 tattgatatt tatatcattt tacgtttctc gttcagcttt cttgtacaaa gtggttgatc
     3181 tagagggccc gcagtggggt cagcttctac cggggcggcg tcccggggtc ttggaactgc
     3241 atgcagactg cctgcttggg aaacatactt ctttatatgc ccatatggac ctgctaagct
     3301 atggaatgta aagaagtatg tatctcaggc cgggacctct ctcgccgcac tgaggggcac
     3361 tccacaccac gggggccgcc gcggccgaca gccagccagg cggctgtggt gggggtgagg
     3421 ctggggcggc aggctgcctg ggcgttggtg gacggtcgtg tccccgccag cattgttctg
     3481 tcactttgcc tgtgtcagca ggtggaagtg ctcaccagac attcgaaggt aagcctatcc
     3541 ctaaccctct cctcggtctc gattctacgc gtaccggtta gtaatgagtt taaacggggg
     3601 aggctaactg aaacacggaa ggagacaata ccggaaggaa cccgcgctat gacggcaata
     3661 aaaagacaga ataaaacgca cgggtgttgg gtcgtttgtt cataaacgcg gggttcggtc
     3721 ccagggctgg cactctgtcg ataccccacc gagaccccat tggggccaat acgcccgcgt
     3781 ttcttccttt tccccacccc accccccaag ttcgggtgaa ggcccagggc tcgcagccaa
     3841 cgtcggggcg gcaggccctg ccatagcaga tctgcgcagc tggggctcta gggggtatcc
     3901 ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     3961 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     4021 cacgttcgcc ggctttcccc gtcaagctct aaatcggggc atccctttag ggttccgatt
     4081 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
     4141 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     4201 tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt
     4261 ataagggatt ttggggattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
     4321 taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
     4381 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
     4441 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     4501 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     4561 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct
     4621 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc
     4681 ccgggagctt gtatatccat tttcggatct gatcaagaga caggatgagg atcgtttcgc
     4741 atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc
     4801 ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca
     4861 gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg
     4921 caggacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg
     4981 ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag
     5041 gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg
     5101 cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc
     5161 atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa
     5221 gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac
     5281 ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat
     5341 ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac
     5401 atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc
     5461 ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt
     5521 gacgagttct tctgagcggg actctggggt tcgcgaaatg accgaccaag cgacgcccaa
     5581 cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat
     5641 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt
     5701 cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     5761 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     5821 caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg
     5881 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     5941 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     6001 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     6061 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     6121 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     6181 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     6241 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     6301 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     6361 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     6421 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcaatg
     6481 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     6541 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     6601 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     6661 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     6721 aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     6781 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     6841 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     6901 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     6961 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     7021 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     7081 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     7141 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     7201 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     7261 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     7321 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     7381 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     7441 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     7501 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     7561 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     7621 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     7681 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     7741 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     7801 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     7861 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     7921 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     7981 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtc
//
