LOCUS       sequence-223560-        4020 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Entry clone containing AtMIR390a backbone (split by ccdB gene). Used
            to make artificial microRNA construct. For use in plants and 
            compatible with the MultiSite Gateway system..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4020)
  AUTHORS   Wang X, Ye L, Lyu M, Ursache R, Loytynoja A, Mahonen AP
  TITLE     An inducible genome editing system for plants.
  JOURNAL   Nat Plants. 2020 Jun 29. pii: 10.1038/s41477-020-0695-2. doi: 
            10.1038/s41477-020-0695-2.
  PUBMED    32601420
REFERENCE   2  (bases 1 to 4020)
  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: "Nat Plants.
            2020 Jun 29. pii: 10.1038/s41477-020-0695-2. doi: 
            10.1038/s41477-020-0695-2."
COMMENT     Sequence Label: p221z-AtMIR390a
FEATURES             Location/Qualifiers
     source          1..4020
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     522..544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     536..553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    569..668
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(962..967,968..985,986..992)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             1046..1705
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1112..1131)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             2047..2352
                     /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     2231..2250
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(2616..2715)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2732..2751)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2733..2751)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2756..2772)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2756..2772)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             complement(2840..3214)
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     promoter        complement(3233..3280)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     rep_origin      3370..3958
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3859..3878
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggcccca aataatgatt ttattttgac tgatagtgac
      601 ctgttcgttg caacacattg atgagcaatg cttttttata atgccaactt tgtacaaaaa
      661 agcaggctcc gcggccgccc ccttcaccta taggggggaa aaaaaggtag tcatcagata
      721 tatattttgg taagaaaata tagaaatgaa taatttcacg tttaacgaag aggagatgac
      781 gtgtgttcct tcgaacccga gttttgttcg tctataaata gcaccttctc ttctccttct
      841 tcctcacttc catcttttta gcttcactat ctctctataa tcggttttat ctttctctaa
      901 gtcacaaccc aaaaaaacaa agtagagaag aatctgtaag agaccattag gcaccccagg
      961 ctttacactt tatgcttccg gctcgtataa tgtgtggatt ttgagttagg agccgtcgag
     1021 attttcagga gctaaggaag ctaaaatgga gaaaaaaatc actggatata ccaccgttga
     1081 tatatcccaa tggcatcgta aagaacattt tgaggcattt cagtcagttg ctcaatgtac
     1141 ctataaccag accgttcagc tggatattac ggccttttta aagaccgtaa agaaaaataa
     1201 gcacaagttt tatccggcct ttattcacat tcttgcccgc ctgatgaatg ctcatccgga
     1261 gttccgtatg gcaatgaaag acggtgagct ggtgatatgg gatagtgttc acccttgtta
     1321 caccgttttc catgagcaaa ctgaaacgtt ttcatcgctc tggagtgaat accacgacga
     1381 tttccggcag tttctacaca tatattcgca agatgtggcg tgttacggtg aaaacctggc
     1441 ctatttccct aaagggttta ttgagaatat gtttttcgtc tcagccaatc cctgggtgag
     1501 tttcaccagt tttgatttaa acgtggccaa tatggacaac ttcttcgccc ccgttttcac
     1561 catgggcaaa tattatacgc aaggcgacaa ggtgctgatg ccgctggcga ttcaggttca
     1621 tcatgccgtt tgtgatggct tccatgtcgg cagaatgctt aatgaattac aacagtactg
     1681 cgatgagtgg cagggcgggg cgtaaacgcg tggagccggc ttactaaaag ccagataaca
     1741 gtatgcgtat ttgcgcgctg atttttgcgg tataagaata tatactgata tgtatacccg
     1801 aagtatgtca aaaagaggta tgctatgaag cagcgtatta cagtgacagt tgacagcgac
     1861 agctatcagt tgctcaaggc atatatgatg tcaatatctc cggtctggta agcacaacca
     1921 tgcagaatga agcccgtcgt ctgcgtgccg aacgctggaa agcggaaaat caggaaggga
     1981 tggctgaggt cgcccggttt attgaaatga acggctcttt tgctgacgag aacaggggct
     2041 ggtgaaatgc agtttaaggt ttacacctat aaaagagaga gccgttatcg tctgtttgtg
     2101 gatgtacaga gtgatattat tgacacgccc ggccgacgga tggtgatccc cctggccagt
     2161 gcacgtctgc tgtcagataa agtctcccgt gaactttacc cggtggtgca tatcggggat
     2221 gaaagctggc gcatgatgac caccgatatg gccagtgtgc cggtttccgt tatcggggaa
     2281 gaagtggctg atctcagcca ccgcgaaaat gacatcaaaa acgccattaa cctgatgttc
     2341 tggggaatat aaatgtcagg ctcccttata cacagccagt ctgcacctcg acggtctcac
     2401 attggctctt cttactacaa tgaaaaaggc cgaggcaaaa cgcctaaaat cacttgagaa
     2461 tcaattcttt ttactgtcca tttaagctat cttttataaa cgtgtcttat tttctatctc
     2521 ttttgtttaa actaagaaac tatagtattt tgtctaaaac aaaacatgaa agaacagatt
     2581 agatctcatc tttagtctca agggtgggcg cgccgaccca gctttcttgt acaaagttgg
     2641 cattataaga aagcattgct tatcaatttg ttgcaacgaa caggtcacta tcagtcaaaa
     2701 taaaatcatt atttgccatc cagctgatat cccctatagt gagtcgtatt acatggtcat
     2761 agctgtttcc tggcagctct ggcccgtgtc tcaaaatctc tgatgttaca ttgcacaaga
     2821 taaaataata tcatcatgat cagtcctgct cctcggccac gaagtgcacg cagttgccgg
     2881 ccgggtcgcg cagggcgaac tcccgccccc acggctgctc gccgatctcg gtcatggccg
     2941 gcccggaggc gtcccggaag ttcgtggaca cgacctccga ccactcggcg tacagctcgt
     3001 ccaggccgcg cacccacacc caggccaggg tgttgtccgg caccacctgg tcctggaccg
     3061 cgctgatgaa cagggtcacg tcgtcccgga ccacaccggc gaagtcgtcc tccacgaagt
     3121 cccgggagaa cccgagccgg tcggtccaga actcgaccgc tccggcgacg tcgcgcgcgg
     3181 tgagcaccgg aacggcactg gtcaacttgg ccatggttta gttcctcacc ttgtcgtatt
     3241 atactatgcc gatatactat gccgatgatt aattgtcaac acgtgctgat catgaccaaa
     3301 atcccttaac gtgagttacg cgtcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     3361 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     3421 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     3481 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg
     3541 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     3601 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     3661 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     3721 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     3781 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     3841 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     3901 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     3961 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
//
