LOCUS       sequence-475345-        3704 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  M13 phagemid constitutively expressing sgRNA-B0. sgRNA-B0 is a dummy
            20-nt sequence, containing a Golden Gate cloning site (BsaI). (A 
            gift of the Jaramillo Lab, where it is called PAJ552.).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3704)
  AUTHORS   Kushwaha M, Singh V, Jaramillo A
  TITLE     Development of a genetically-encoded in vivo computing platform to 
            play the game of Tic-Tac-Toe
  JOURNAL   Zenodo (2017) Conference: SB7.0: The Seventh International Meeting 
            on Synthetic Biology.
REFERENCE   2  (bases 1 to 3704)
  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: "Zenodo 
            (2017) Conference: SB7.0: The Seventh International Meeting on 
            Synthetic Biology."
COMMENT     Sequence Label: pSEVA63_sgRNA-B0_M13ps
FEATURES             Location/Qualifiers
     source          1..3704
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(15..524)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(210..231)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     418..437
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(627..713)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     728..750
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     promoter        781..815
                     /label=J23119 promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119)"
     misc_RNA        836..911
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     terminator      976..1070
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     promoter        join(1101..1106,1107..1123,1124..1129)
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     CDS             1318..1851
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     oriT            2062..2170
                     /note="incP origin of transfer"
     CDS             complement(2181..2843)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     rep_origin      2844..3615
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
ORIGIN
        1 cgcgagattg atttaccccg gttgataatc agaaaagccc caaaaacagg aagattgtat
       61 aagcaaatat ttaaattgta aacgttaata ttttgttaaa attcgcgtta aatttttgtt
      121 aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag
      181 aatagcccga gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga
      241 acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg
      301 aaccatcacc caaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc
      361 ctaaagggag cccccgattt agagcttgac ggggaaagcg aacgtggcga gaaaggaagg
      421 gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt
      481 aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtaaaagg atctaggtga
      541 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
      601 gcgcgcccag ctgtctaggg cggcggattt gtcctactca ggagagcgtt caccgacaaa
      661 caacagataa aacgaaaggc ccagtctttc gactgagcct ttcgttttat ttgatgcctt
      721 taattaaagc ggataacaat ttcacacagg aggccgccta ggccgcggcc gcgcgaattc
      781 ttgacagcta gctcagtcct aggtataatg ctagcagaga ccactttcgg tctctgtttt
      841 agagctagaa atagcaagtt aaaataaggc tagtccgtta tcaacttgaa aaagtggcac
      901 cgagtcggtg cttttctgca ggcatgcaag cttgcggccg cgtcgtgact gggaaaaccc
      961 tggcgactag tcttggactc ctgttgatag atccagtaat gacctcagaa ctccatctgg
     1021 atttgttcag aacgctcggt tgccgccggg cgttttttat tggtgagaat ccaggggtcc
     1081 ccaataatta cgatttaaat ttgacataag cctgttcggt tcgtaaactg taatgcaagt
     1141 agcgtatgcg ctcacgcaac tggtccagaa ccttgaccga acgcagcggt ggtaacggcg
     1201 cagtggcggt tttcatggct tgttatgact gtttttttgt acagcctatg cctcgggcat
     1261 ccaagcagca agcgcgttac gccgtgggtc gatgtttgat gttatggagc agcaacgatg
     1321 ttacgcagca gcaacgatgt tacgcagcag ggcagtcgcc ctaaaacaaa gttaggtggc
     1381 tcaagtatgg gcatcattcg cacatgtagg ctcggccctg accaagtcaa atccatgcgg
     1441 gctgctcttg atcttttcgg tcgtgagttc ggagacgtag ccacctactc ccaacatcag
     1501 ccggactccg attacctcgg gaacttgctc cgtagtaaga cattcatcgc gcttgctgcc
     1561 ttcgaccaag aagcggttgt tggcgctctc gcggcttacg ttctgcccaa gtttgagcag
     1621 ccgcgtagtg agatctatat ctatgatctc gcagtctccg gagagcaccg gaggcagggc
     1681 attgccaccg cgctcatcaa tctcctcaag catgaggcca acgcgcttgg tgcttatgtg
     1741 atctacgtgc aagcagatta cggtgacgat cccgcagtgg ctctctatac aaagttgggc
     1801 atacgggaag aagtgatgca ctttgatatc gacccaagta ccgccaccta acaattcgtt
     1861 caagccgaga tcggcttccc ggccgcggag ttgttcggta aattggacaa cggtccgcgc
     1921 gttgtccttt tccgctgcat aaccctgctt cggggtcatt atagcgattt tttcggtata
     1981 tccatccttt ttcgcacgat atacaggatt ttgccaaagg gttcgtgtag actttccttg
     2041 gtgtatccaa cggcgtcagc cgggcaggat aggtgaagta ggcccacccg cgagcgggtg
     2101 ttccttcttc actgtccctt attcgcacct ggcggtgctc aacgggaatc ctgctctgcg
     2161 aggctggccg taggccggcc ctaccggcgc ggcagcgtta cccgtgtcgg cggctccaac
     2221 ggctcgccat cgtccagaaa acacggctca tcgggcatcg gcaggcgctg ctgcccgcgc
     2281 cgttcccatt cctccgtttc ggtcaaggct ggcaggtctg gttccatgcc cggaatgccg
     2341 ggctggctgg gcggctcctc gccggggccg gtcggtagtt gctgctcgcc cggatacagg
     2401 gtcgggatgc ggcgcaggtc gccatgcccc aacagcgatt cgtcctggtc gtcgtgatca
     2461 accaccacgg cggcactgaa caccgacagg cgcaactggt cgcggggctg gccccacgcc
     2521 acgcggtcat tgaccacgta ggccgacacg gtgccggggc cgttgagctt cacgacggag
     2581 atccagcgct cggccaccaa gtccttgact gcgtattgga ccgtccgcaa agaacgtccg
     2641 atgagcttgg aaagtgtctt ctggctgacc accacggcgt tctggtggcc catctgcgcc
     2701 acgaggtgat gcagcagcat tgccgccgtg ggtttcctcg caataagccc ggcccacgcc
     2761 tcatgcgctt tgcgttccgt ttgcacccag tgaccgggct tgttcttggc ttgaatgccg
     2821 atttctctgg actgcgtggc catgcttatc tccatgcggt aggggtgccg cacggttgcg
     2881 gcaccatgcg caatcagctg caacttttcg gcagcgcgac aacaattatg cgttgcgtaa
     2941 aagtggcagt caattacaga ttttctttaa cctacgcaat gagctattgc ggggggtgcc
     3001 gcaatgagct gttgcgtacc cccctttttt aagttgttga tttttaagtc tttcgcattt
     3061 cgccctatat ctagttcttt ggtgcccaaa gaagggcacc cctgcggggt tcccccacgc
     3121 cttcggcgcg gctccccctc cggcaaaaag tggcccctcc ggggcttgtt gatcgactgc
     3181 gcggccttcg gccttgccca aggtggcgct gcccccttgg aacccccgca ctcgccgccg
     3241 tgaggctcgg ggggcaggcg ggcgggcttc gcccttcgac tgcccccact cgcataggct
     3301 tgggtcgttc caggcgcgtc aaggccaagc cgctgcgcgg tcgctgcgcg agccttgacc
     3361 cgccttccac ttggtgtcca accggcaagc gaagcgcgca ggccgcaggc cggaggcttt
     3421 tccccagaga aaattaaaaa aattgatggg gcaaggccgc aggccgcgca gttggagccg
     3481 gtgggtatgt ggtcgaaggc tgggtagccg gtgggcaatc cctgtggtca agctcgtggg
     3541 caggcgcagc ctgtccatca gcttgtccag cagggttgtc cacgggccga gcgaagcgag
     3601 ccagccggtg gccgctcgcg gccatcgtcc acatatccac gggctggcaa gggagcgcag
     3661 cgaccgcgca gggcgaagcc cggagagcaa gcccgtaggg gggg
//
