LOCUS       sequence-79410-e        7916 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plant expression vector (35S) for direct cloning of artificial 
            microRNAs into Arabidopsis thaliana MIR390a precursor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7916)
  AUTHORS   Carbonell A, Takeda A, Fahlgren N, Johnson SC, Cuperus JT, 
            Carrington JC
  TITLE     New Generation of Artificial MicroRNA and Synthetic Trans-Acting 
            Small Interfering RNA Vectors for Efficient Gene Silencing in 
            Arabidopsis.
  JOURNAL   Plant Physiol. 2014 Mar 19.
  PUBMED    24647477
REFERENCE   2  (bases 1 to 7916)
  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: "Plant 
            Physiol. 2014 Mar 19."
COMMENT     Sequence Label: pFK210B-AtMIR390a-B/c
FEATURES             Location/Qualifiers
     source          1..7916
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    1..23
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA 
                     (truncated)"
     terminator      complement(47..299)
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     CDS             complement(319..870)
                     /codon_start=1
                     /gene="Streptomyces hygroscopicus bar"
                     /product="phosphinothricin acetyltransferase"
                     /label=BlpR
                     /note="confers resistance to bialophos or phosphinothricin"
                     /translation="MSPERRPADIRRATEADMPAVCTIVNHYIETSTVNFRTEPQEPQE
                     WTDDLVRLRERYPWLVAEVDGEVAGIAYAGPWKARNAYDWTAESTVYVSPRHQRTGLGS
                     TLYTHLLKSLEAQGFKSVVAVIGLPNDPSVRMHEALGYAPRGMLRAAGFKHGNWHDVGF
                     WQLDFSLPVPPRPVLPVTEI"
     promoter        complement(912..1091)
                     /label=NOS promoter
                     /note="nopaline synthase promoter"
     primer_bind     1328..1350
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1342..1359
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1343..1359
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1369..1388
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1369..1387
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     1412..1428
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     1413..1429
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        1966..2311
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     2254..2274
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     protein_bind    2336..2360
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     promoter        join(2654..2659,2660..2677,2678..2684)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             2738..3397
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2804..2823)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             3739..4044
                     /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     3923..3942
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(4308..4332)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     terminator      4339..4980
                     /label=E9 terminator
                     /note="terminator and polyadenylation signal from the pea 
                     rbcS-E9 gene"
     promoter        complement(5002..5020)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5041..5057)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5041..5057)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5054..5076)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5065..5081
                     /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(5089..5095,5096..5113,5114..5119))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5134..5155
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5272..5289)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5394..5418
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     rep_origin      complement(5509..6097)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5589..5608)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6268..7056)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      7347..7782
                     /label=pSa ori
                     /note="origin of replication from bacterial plasmid pSa"
ORIGIN
        1 tggcaggata tattgtggtg taacgttatc agcttgcatg ccggtcgatc tagtaacata
       61 gatgacaccg cgcgcgataa tttatcctag tttgcgcgct atattttgtt ttctatcgcg
      121 tattaaatgt ataattgcgg gactctaatc ataaaaaccc atctcataaa taacgtcatg
      181 cattacatgt taattattac atgcttaacg taattcaaca gaaattatat gataatcatc
      241 gcaagaccgg caacaggatt caatcttaag aaactttatt gccaaatgtt tgaacgatct
      301 gcttgactct aggggtcatc agatttcggt gacgggcagg accggacggg gcggcaccgg
      361 caggctgaag tccagctgcc agaaacccac gtcatgccag ttcccgtgct tgaagccggc
      421 cgcccgcagc atgccgcggg gggcatatcc gagcgcctcg tgcatgcgca cgctcgggtc
      481 gttgggcagc ccgatgacag cgaccacgct cttgaagccc tgtgcctcca gggacttcag
      541 caggtgggtg tagagcgtgg agcccagtcc cgtccgctgg tggcgggggg agacgtacac
      601 ggtggactcg gccgtccagt cgtaggcgtt gcgtgccttc cagggacccg cgtaggcgat
      661 gccggcgacc tcgccgtcca cctcggcgac gagccaggga tagcgctccc gcagacggac
      721 gaggtcgtcc gtccactcct gcggttcctg cggctcggta cggaagttga ccgtgcttgt
      781 ctcgatgtag tggttgacga tggtgcagac cgccggcatg tccgcctcgg tggcacggcg
      841 gatgtcggcc gggcgtcgtt ctgggctcat ggtagatccc ctcgatcgag ttgagagtga
      901 atatgagact ctaattggat accgagggga atttatggaa cgtcagtgga gcatttttga
      961 caagaaatat ttgctagctg atagtgacct taggcgactt ttgaacgcgc aataatggtt
     1021 tctgacgtat gtgcttagct cattaaactc cagaaacccg cggctcagtg gctccttcaa
     1081 cgttgcggtt ctgtcagttc caaacgtaaa acggcttgtc ccgcgtcatc ggcgggggtc
     1141 ataacgtgac tcccttaatt ctccgctcat gtatcgataa cattaacgtt tacaatttcg
     1201 cgccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg
     1261 ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg ggtaacgcca
     1321 gggttttccc agtcacgacg ttgtaaaacg acggccagtg agcgcgcgta atacgactca
     1381 ctatagggcg aattgggtac cgggcccccc ctcgaggtcg acggtatcga taagcttgat
     1441 atcgaattcc tgcagcccgg gggatccatt cggtccccag attagccttt tcaatttcag
     1501 aaagaatgct aacccacaga tggttagaga ggcttacgca gcaggtttca tcaagacgat
     1561 ctacccgagc aataatctcc aggaaatcaa ataccttccc aagaaggtta aagatgcagt
     1621 caaaagattc aggactaact gcatcaagaa cacagagaaa gatatatttc tcaagatcag
     1681 aagtactatt ccagtatgga cgattcaagg cttgcttcac aaaccaaggc aagtaataga
     1741 gattggagtc tctaaaaagg tagttcccac tgaatcaaag gccatggagt caaagattca
     1801 aatagaggac ctaacagaac tcgccgtaaa gactggcgaa cagttcatac agagtctctt
     1861 acgactcaat gacaagaaga aaatcttcgt caacatggtg gagcacgaca cacttgtcta
     1921 ctccaaaaat atcaaagata cagtctcaga agaccaaagg gcaattgaga cttttcaaca
     1981 aagggtaata tccggaaacc tcctcggatt ccattgccca gctatctgtc actttattgt
     2041 gaagatagtg gaaaaggaag gtggctccta caaatgccat cattgcgata aaggaaaggc
     2101 catcgttgaa gatgcctctg ccgacagtgg tcccaaagat ggacccccac ccacgaggag
     2161 catcgtggaa aaagaagacg ttccaaccac gtcttcaaag caagtggatt gatgtgatat
     2221 ctccactgac gtaagggatg acgcacaatc ccactatcct tcgcaagacc cttcctctat
     2281 ataaggaagt tcatttcatt tggagagaac acgggggacg agcttctaga ggatcacaag
     2341 tttgtacaaa aaagcaggct ccgcggccgc ccccttcacc tatagggggg aaaaaaaggt
     2401 agtcatcaga tatatatttt ggtaagaaaa tatagaaatg aataatttca cgtttaacga
     2461 agaggagatg acgtgtgttc cttcgaaccc gagttttgtt cgtctataaa tagcaccttc
     2521 tcttctcctt cttcctcact tccatctttt tagcttcact atctctctat aatcggtttt
     2581 atctttctct aagtcacaac ccaaaaaaac aaagtagaga agaatctgta agagaccatt
     2641 aggcacccca ggctttacac tttatgcttc cggctcgtat aatgtgtgga ttttgagtta
     2701 ggagccgtcg agattttcag gagctaagga agctaaaatg gagaaaaaaa tcactggata
     2761 taccaccgtt gatatatccc aatggcatcg taaagaacat tttgaggcat ttcagtcagt
     2821 tgctcaatgt acctataacc agaccgttca gctggatatt acggcctttt taaagaccgt
     2881 aaagaaaaat aagcacaagt tttatccggc ctttattcac attcttgccc gcctgatgaa
     2941 tgctcatccg gagttccgta tggcaatgaa agacggtgag ctggtgatat gggatagtgt
     3001 tcacccttgt tacaccgttt tccatgagca aactgaaacg ttttcatcgc tctggagtga
     3061 ataccacgac gatttccggc agtttctaca catatattcg caagatgtgg cgtgttacgg
     3121 tgaaaacctg gcctatttcc ctaaagggtt tattgagaat atgtttttcg tctcagccaa
     3181 tccctgggtg agtttcacca gttttgattt aaacgtggcc aatatggaca acttcttcgc
     3241 ccccgttttc accatgggca aatattatac gcaaggcgac aaggtgctga tgccgctggc
     3301 gattcaggtt catcatgccg tttgtgatgg cttccatgtc ggcagaatgc ttaatgaatt
     3361 acaacagtac tgcgatgagt ggcagggcgg ggcgtaaacg cgtggagccg gcttactaaa
     3421 agccagataa cagtatgcgt atttgcgcgc tgatttttgc ggtataagaa tatatactga
     3481 tatgtatacc cgaagtatgt caaaaagagg tatgctatga agcagcgtat tacagtgaca
     3541 gttgacagcg acagctatca gttgctcaag gcatatatga tgtcaatatc tccggtctgg
     3601 taagcacaac catgcagaat gaagcccgtc gtctgcgtgc cgaacgctgg aaagcggaaa
     3661 atcaggaagg gatggctgag gtcgcccggt ttattgaaat gaacggctct tttgctgacg
     3721 agaacagggg ctggtgaaat gcagtttaag gtttacacct ataaaagaga gagccgttat
     3781 cgtctgtttg tggatgtaca gagtgatatt attgacacgc ccggccgacg gatggtgatc
     3841 cccctggcca gtgcacgtct gctgtcagat aaagtctccc gtgaacttta cccggtggtg
     3901 catatcgggg atgaaagctg gcgcatgatg accaccgata tggccagtgt gccggtttcc
     3961 gttatcgggg aagaagtggc tgatctcagc caccgcgaaa atgacatcaa aaacgccatt
     4021 aacctgatgt tctggggaat ataaatgtca ggctccctta tacacagcca gtctgcacct
     4081 cgacggtctc acattggctc ttcttactac aatgaaaaag gccgaggcaa aacgcctaaa
     4141 atcacttgag aatcaattct ttttactgtc catttaagct atcttttata aacgtgtctt
     4201 attttctatc tcttttgttt aaactaagaa actatagtat tttgtctaaa acaaaacatg
     4261 aaagaacaga ttagatctca tctttagtct caagggtggg cgcgccgacc cagctttctt
     4321 gtacaaagtg gtgatcctag ctttcgttcg tatcatcggt ttcgacaacg ttcgtcaagt
     4381 tcaatgcatc agtttcattg cgcacacacc agaatcctac tgagtttgag tattatggca
     4441 ttgggaaaac tgtttttctt gtaccatttg ttgtgcttgt aatttactgt gttttttatt
     4501 cggttttcgc tatcgaactg tgaaatggaa atggatggag aagagttaat gaatgatatg
     4561 gtccttttgt tcattctcaa attaatatta tttgtttttt ctcttatttg ttgtgtgttg
     4621 aatttgaaat tataagagat atgcaaacat tttgttttga gtaaaaatgt gtcaaatcgt
     4681 ggcctctaat gaccgaagtt aatatgagga gtaaaacact tgtagttgta ccattatgct
     4741 tattcactag gcaacaaata tattttcaga cctagaaaag ctgcaaatgt tactgaatac
     4801 aagtatgtcc tcttgtgttt tagacattta tgaactttcc tttatgtaat tttccagaat
     4861 ccttgtcaga ttctaatcat tgctttataa ttatagttat actcatggat ttgtagttga
     4921 gtatgaaaat attttttaat gcattttatg acttgccaat tgattgacaa catgcatcaa
     4981 tcggagctcc agcttttgtt ccctttagtg agggttaatt ccgagcttgg cgtaatcatg
     5041 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     5101 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     5161 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     5221 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     5281 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     5341 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgaagg ccttgacagg
     5401 atatattggc gggtaaacta agtcgctgta tgtgtttgtt tgagatctca tgtgagcaaa
     5461 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     5521 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     5581 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     5641 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     5701 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     5761 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     5821 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     5881 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     5941 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaagaag
     6001 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     6061 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     6121 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     6181 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     6241 tatatatgtg taacattggt ctagtgatta tttgccgact accttggtga tctcgccttt
     6301 cacgtagtga acaaattctt ccaactgatc tgcgcgcgag gccaagcgat cttcttgtcc
     6361 aagataagcc tgcctagctt caagtatgac gggctgatac tgggccggca ggcgctccat
     6421 tgcccagtcg gcagcgacat ccttcggcgc gattttgccg gttactgcgc tgtaccaaat
     6481 gcgggacaac gtaagcacta catttcgctc atcgccagcc cagtcgggcg gcgagttcca
     6541 tagcgttaag gtttcattta gcgcctcaaa tagatcctgt tcaggaaccg gatcaaagag
     6601 ttcctccgcc gctggaccta ccaaggcaac gctatgttct cttgcttttg tcagcaagat
     6661 agccagatca atgtcgatcg tggctggctc gaagatacct gcaagaatgt cattgcgctg
     6721 ccattctcca aattgcagtt cgcgcttagc tggataacgc cacggaatga tgtcgtcgtg
     6781 cacaacaatg gtgacttcta cagcgcggag aatctcgctc tctccagggg aagccgaagt
     6841 ttccaaaagg tcgttgatca aagctcgccg cgttgtttca tcaagcctta cggtcaccgt
     6901 aaccagcaaa tcaatatcac tgtgtggctt caggccgcca tccactgcgg agccgtacaa
     6961 atgtacggcc agcaacgtcg gttcgagatg gcgctcgatg acgccaacta cctctgatag
     7021 ttgagtcgat acttcggcga tcaccgcttc cctcataaca ccccttgtat tactgtttat
     7081 gtaagcagac agttttattg ttcatgatga tatattttta tcttgtgcaa tgtaacatca
     7141 gagattttga gacacaacgt ggctttgttg aataaatcga acttttgctg agttgaagga
     7201 tcagatcacg catcttcccg acaacgcaga ccgttccgtg gcaaagcaaa agttcaaaat
     7261 caccaactgg tccacctaca acaaagctct catcaaccgt ggctccctca ctttctggct
     7321 ggatgatggg gcgattcagg cgatccccat ccaacagccc gccgtcgagc gggctttttt
     7381 atccccggaa gcctgtggat agagggtagt tatccacgtg aaaccgctaa tgccccgcaa
     7441 agccttgatt cacggggctt tccggcccgc tccaaaaact atccacgtga aatcgctaat
     7501 cagggtacgt gaaatcgcta atcggagtac gtgaaatcgc taataaggtc acgtgaaatc
     7561 gctaatcaaa aaggcacgtg agaacgctaa tagccctttc agatcaacag cttgcaaaca
     7621 cccctcgctc cggcaagtag ttacagcaag tagtatgttc aattagcttt tcaattatga
     7681 atatatatat caattattgg tcgcccttgg cttgtggaca atgcgctacg cgcaccggct
     7741 ccgcccgtgg acaaccgcaa gcggttgccc accgtcgagc gccagcgcct ttgcccacaa
     7801 cccggcggcc ggccgcaaca gatcgtttta taaatttttt tttttgaaaa agaaaaagcc
     7861 cgaaaggcgg caacctctcg ggcttctgga tttccgatcc ccggaattag agatct
//
