LOCUS       sequence-348326-        2713 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  First sgRNA in a tRNA separated array of sgRNAs.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2713)
  AUTHORS   Cody JP, Maher MF, Nasti RA, Starker CG, Chamness JC, Voytas DF
  TITLE     Direct delivery and fast-treated Agrobacterium co-culture 
            (Fast-TrACC) plant transformation methods for Nicotiana benthamiana.
  JOURNAL   Nat Protoc. 2022 Oct 17. pii: 10.1038/s41596-022-00749-9. doi: 
            10.1038/s41596-022-00749-9.
  PUBMED    36253612
REFERENCE   2  (bases 1 to 2713)
  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 Protoc.
            2022 Oct 17. pii: 10.1038/s41596-022-00749-9. doi: 
            10.1038/s41596-022-00749-9."
COMMENT     Sequence Label: pTAG5
FEATURES             Location/Qualifiers
     source          1..2713
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    195..216
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(231..236,237..254,255..261)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    269..285
                     /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     274..296
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     293..309
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     293..309
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             305..646
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL
                     QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGATSRLR
                     FRARNSKLK"
     primer_bind     328..348
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        330..348
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(395..414)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(396..414)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(424..441)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(424..440)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(433..455)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             952..1743
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MGEAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      2084..2672
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2573..2592
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ggtctcagca agttaaaata aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc
       61 ggtgcaacaa agcaccagtg gtctagtggt agaatagtac cctgccacgg tacagacccg
      121 ggttcgattc ccggctggtg catgagacga gatctagacg accggtggaa agcgggcagt
      181 gagcgcaacg caattaatgt gagttagctc actcattagg caccccaggc tttacacttt
      241 atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca cacaggaaac
      301 agctatgacc atgattacgc caagcgcgca attaaccctc actaaaggga acaaaagctg
      361 gaggacctcg agggggggcc cggtacccaa ttcgccctat agtgagtcgt attacgcgcg
      421 ctcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa
      481 tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga
      541 tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg gacgcgccct gtagcggcgc
      601 cactagtcgt ctccgtttta gagctagaaa tagcaagtta aaataaggct agtcccgaga
      661 ccacgaagtg ccgaggcacg aacccagtgg acataagcct gttcggttcg taagctgtaa
      721 tgcaagtagc gtatgcgctc acgcaactgg tccagaacct tgaccgaacg cagcggtggt
      781 aacggcgcag tggcggtttt catggcttgt tatgactgtt tttttggggt acagtctatg
      841 cctcgggcat ccaagcagca agcgcgttac gccgtgggtc gatgtttgat gttatggagc
      901 agcaacgatg ttacgcagca gggcagtcgc cctaaaacaa agttaaacat catgggggaa
      961 gcggtgatcg ccgaagtatc gactcaacta tcagaggtag ttggcgtcat cgagcgccat
     1021 ctcgaaccga cgttgctggc cgtacatttg tacggctccg cagtggatgg cggcctgaag
     1081 ccacacagcg atattgattt gctggttacg gtgaccgtaa ggcttgatga aacaacgcgg
     1141 cgagctttga tcaacgacct tttggaaact tcggcttccc ctggagagag cgagattctc
     1201 cgcgctgtag aagtcaccat tgttgtgcac gacgacatca ttccgtggcg ttatccagct
     1261 aagcgcgaac tgcaatttgg agaatggcag cgcaatgaca ttcttgcagg tatcttcgag
     1321 ccagccacga tcgacattga tctggctatc ttgctgacaa aagcaagaga acatagcgtt
     1381 gccttggtag gtccagcggc ggaggaactc tttgatccgg ttcctgaaca ggatctattt
     1441 gaggcgctaa atgaaacctt aacgctatgg aactcgccgc ccgactgggc tggcgatgag
     1501 cgaaatgtag tgcttacgtt gtcccgcatt tggtacagcg cagtaaccgg caaaatcgcg
     1561 ccgaaggatg tcgctgccga ctgggcaatg gagcgcctgc cggcccagta tcagcccgtc
     1621 atacttgaag ctagacaggc ttatcttgga caagaagaag atcgcttggc ctcgcgcgca
     1681 gatcagttgg aagaatttgt ccattacgtg aaaggcgaga tcaccaaggt agtcggcaaa
     1741 taatgtctag ctagaaattc gttcaagccg acgccgcttc gcggcgcggc ttaactcaag
     1801 cgttagatgc actaagcaca taattgctca cagccaaact atcaggtcaa gtctgctttt
     1861 attattttta agcgtgcata ataagcccta cacaaattgg gagatatatc atgctgtcag
     1921 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     1981 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     2041 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     2101 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     2161 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     2221 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     2281 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     2341 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     2401 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     2461 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     2521 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     2581 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     2641 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     2701 tcctgcactc tgt
//
