LOCUS       sequence-405609-        3347 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Golden Gate recipient and Gateway entry vector; assembly of 4 gRNAs 
            .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3347)
  AUTHORS   Lowder LG, Paul JW, Baltes NJ, Voytas DF, Zhang Y, Zhang D, Tang X, 
            Zheng X, Hsieh TF, Qi Y
  TITLE     A CRISPR/Cas9 toolbox for multiplexed plant genome editing and 
            transcriptional regulation.
  JOURNAL   Plant Physiol. 2015 Aug 21. pii: pp.00636.2015.
  PUBMED    26297141
REFERENCE   2  (bases 1 to 3347)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Plant 
            Physiol. 2015 Aug 21. pii: pp.00636.2015."
COMMENT     Sequence Label: pYPQ144
FEATURES             Location/Qualifiers
     source          1..3347
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        22..40
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        57..75
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    93..192
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    257..278
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(293..298,299..316,317..323)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    331..347
                     /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     336..358
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     355..371
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             367..804
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL
                     QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGAGLDSH
                     HLGILRYLEGTPWGNFCIQSWHYKKALLINLLQRTGHYQSK"
     primer_bind     390..410
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        392..410
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        complement(458..476)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(486..502)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(720..815)
                     /gene="mutant version of attL"
                     /label=attL5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(830..846)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     terminator      910..996
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1088..1115
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     complement(1274..1291)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1445..2033)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1525..1544)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2126..2917)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
ORIGIN
        1 caggaaacag ctatgaccat gtaatacgac tcactatagg ggatgctatg accatgtaat
       61 acgactcact ataggggata tcagctggat ggcaaataat gattttattt tgactgatag
      121 tgacctgttc gttgcaacaa attgatgagc aattattttt tataatgcca actttgtaca
      181 agaaagctgg gtcgaattcg cccttactag taacaagaga ccaccggtgg aaagcgggca
      241 gtgagcgcaa cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact
      301 ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa
      361 acagctatga ccatgattac gccaagcgcg caattaaccc tcactaaagg gaacaaaagc
      421 tggaggacct cgaggggggg cccggtaccc aattcgccct atagtgagtc gtattacgcg
      481 cgctcactgg ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt
      541 aatcgccttg cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc
      601 gatcgccctt cccaacagtt gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc
      661 gccggtctcg atagccacca cttggggatc ctgcgttatc tagagggaac gccatggggc
      721 aacttttgta tacaaagttg gcattataaa aaagcattgc tcatcaattt gttgcaacga
      781 acaggtcact atcagtcaaa ataaaatcat tattgggccc gagcttaaga ctggccgtcg
      841 ttttacaacg tcgtgactgg gaaaacatcc atgcaagcgt taacgcgaga gtagggaact
      901 gccaggcatc aaataaaacg aaaggctcag tcggaagact gggcctttcg ttttatctgt
      961 tgtttgtcgg tgaacgctct cctgagtagg acaaatccgc cgggagcgga tttgaacgtt
     1021 gtgaagcaac ggcccggagg gtggcgggca ggacgcccgc cataaactgc caggcatcaa
     1081 actaagcaga aggccatcct gacggatggc ctttttgcgt ttctacaaac tcttcctggc
     1141 tcgcggtacg cgtattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     1201 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     1261 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     1321 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     1381 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     1441 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     1501 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     1561 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     1621 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     1681 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     1741 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     1801 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     1861 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     1921 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     1981 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     2041 ctttgatctt ttctacgggg tctgacgctc agtggaacga cgcgtaactc acgttaaggg
     2101 attttggtca tgggtggctc gagggttatt tgccgactac cttggtgatc tcgcctttca
     2161 cgtagtggac aaattcttcc aactgatctg cgcgcgaggc caagcgatct tcttcttgtc
     2221 caagataagc ctgtctagct tcaagtatga cgggctgata ctgggccggc aggcgctcca
     2281 ttgcccagtc ggcagcgaca tccttcggcg cgattttgcc ggttactgcg ctgtaccaaa
     2341 tgcgggacaa cgtaagcact acatttcgct catcgccagc ccagtcgggc ggcgagttcc
     2401 atagcgttaa ggtttcattt agcgcctcaa atagatcctg ttcaggaacc ggatcaaaga
     2461 gttcctccgc cgctggacct accaaggcaa cgctatgttc tcttgctttt gtcagcaaga
     2521 tagccagatc aatgtcgatc gtggctggct cgaagatacc tgcaagaatg tcattgcgct
     2581 gccattctcc aaattgcagt tcgcgcttag ctggataacg ccacggaatg atgtcgtcgt
     2641 gcacaacaat ggtgacttct acagcgcgga gaatctcgct ctctccaggg gaagccgaag
     2701 tttccaaaag gtcgttgatc aaagctcgcc gcgttgtttc atcaagcctt acggtcaccg
     2761 taaccagcaa atcaatatca ctgtgtggct tcaggccgcc atccactgcg gagccgtaca
     2821 aatgtacggc cagcaacgtc ggttcgagat ggcgctcgat gacgccaact acctctgata
     2881 gttgagtcga tacttcggcg atcaccgctt ccctcataat gtttaacttt gttttagggc
     2941 gactgccctg ctgcgtaaca tcgttgctgc tccataacat caaacatcga cccacggcgt
     3001 aacgcgcttg ctgcttggat gcccgaggca tagactgtac cccaaaaaaa cagtcataac
     3061 aagccatgaa aaccgccact gcgccgttac caccgctgcg ttcggtcaag gttctggacc
     3121 agttgcgtga gcgcatacgc tacttgcatt acagcttacg aaccgaacag gcttatgtcc
     3181 actgggttcg tgccttcatc cgtttccacg gtgtgcgtca cccggcaacc ttgggtagca
     3241 gcgaagtcga ggcatttctg tcctggctgg tccagaggag gcatgatgat atatttttat
     3301 cttgtgcaat gtaacatcag agattttgag acacgggcca gagctgc
//
