LOCUS       sequence-488908-        4882 bp    DNA     circular SYN 09-OCT-2025
DEFINITION  Bacterial expression of mutant T4 Acb1 (H44A,H113A) under control of
            the pJex promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4882)
  AUTHORS   Doherty EE, Adler BA, Yoon PH, Hsieh K, Loi K, Armbuster EG, Lahiri 
            A, Bolling CS, Wilcox XE, Akkati A, Iavarone AT, Pogliano J, Doudna 
            JA
  TITLE     A miniature CRISPR-Cas10 enzyme confers immunity by an inverse 
            signaling pathway.
  JOURNAL   bioRxiv [Preprint]. 2025 Apr 1:2025.03.28.646030. doi: 
            10.1101/2025.03.28.646030.
  PUBMED    40196485
REFERENCE   2  (bases 1 to 4882)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 9, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            [Preprint]. 2025 Apr 1:2025.03.28.646030. doi: 
            10.1101/2025.03.28.646030."
COMMENT     Sequence Label: pBA1802
FEATURES             Location/Qualifiers
     source          1..4882
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      253..475
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             complement(1018..1833)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1018..1833)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     1741..1760
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1825..1844
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     RBS             2602..2610
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     terminator      3432..3503
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3519..3546
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(join(4137..4882,1..205))
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
ORIGIN
        1 caccataagc attttcccta ctgatgttca tcatctgagc gtattggtta taagtgaacg
       61 ataccgtccg ttctttcctt gtagggtttt caatcgtggg gttgagtagt gccacacagc
      121 ataaaattag cttggtttca tgctccgtta agtcatagcg actaatcgct agttcatttg
      181 ctttgaaaac aactaattca gacatacatc tcaattggtc taggtgattt taatcactat
      241 accaattgag atgggctagt caatgataat tactagtcct tttcctttga gttgtgggta
      301 tctgtaaatt ctgctagacc tttgctggaa aacttgtaaa ttctgctata ccctctgtaa
      361 attccgctag acctttgtgt gttttttttg tttatattca agtggttata atttatagaa
      421 taaagaaaga ataaaaaaag ataaaaagaa tagatcccag ccctgtgtat aactcactac
      481 tttagtcagt tccgcagtat tacaaaagga tgtcgcaaac gctgtttgct cctctacaaa
      541 acagacctta aaaccctaaa ggcttaagta gcaccctcgc aagctcgggc aaatcgctga
      601 atattccttt tgtctccgac catcaggcac ctgagtcgct gtctttttcg tgacattcag
      661 ttcgctgcgc tcacggctct ggcagtgaat gggggtaaat ggcactacag gcgcctttta
      721 tggattcatg caaggaaact acccataata caagaaaagc ccgtcacggg cttctcaggg
      781 cgttttatgg cgggtctgct atgtggtgct atctgacttt ttgctgttca gcagttcctg
      841 ccctctgatt ttccagtctg accacttcgg attatcccgt gacaggtcat tcagactggc
      901 taatgcaccc agtaaggcag cggtatcatc aacaggctta cccgtcttac tgtccctgtg
      961 tcagatttct tactttattc aggcgattac gtgttacaac caattaacca attctgatta
     1021 gaaaaactca tcgagcatca aatgaaactg caatttattc atatcaggat tatcaatacc
     1081 atatttttga aaaagccgtt tctgtaatga aggagaaaac tcaccgaggc agttccatag
     1141 gatggcaaga tcctggtatc ggtctgcgat tccgactcgt ccaacatcaa tacaacctat
     1201 taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa tcaccatgag tgacgactga
     1261 atccggtgag aatggcaaaa gcttatgcat ttctttccag acttgttcaa caggccagcc
     1321 attacgctcg tcatcaaaat cactcgcatc aaccaaaccg ttattcattc gtgattgcgc
     1381 ctgagcgagg cgaaatacgc gatcgctgtt aaaaggacaa ttacaaacag gaatcgaatg
     1441 caaccggcgc aggaacactg ccagcgcatc aacaatattt tcacctgaat caggatattc
     1501 ttctaatacc tggaatgctg ttttcccggg gatcgcagtg gtgagtaacc atgcatcatc
     1561 aggagtacgg ataaaatgct tgatggtcgg aagaggcata aattccgtca gccagtttag
     1621 tctgaccatc tcatctgtaa catcattggc aacgctacct ttgccatgtt tcagaaacaa
     1681 ctctggcgca tcgggcttcc catacaatcg atagattgtc gcacctgatt gcccgacatt
     1741 atcgcgagcc catttatacc catataaatc agcatccatg ttggaattta atcgcggcct
     1801 ggagcaagac gtttcccgtt gaatatggct cataacaccc cttgtattac tgtttatgta
     1861 agcagacagt tttattgttc atgatgatat atttttatct tgtgcaatgt aacatcagag
     1921 attttgagac acaacgtggc tttgttgaat aaatcgaact tttgctgagt tgaaggatca
     1981 gcacctgccc gaaccagaga cttatctcag gacgtcttac gaaaataact caagctgaat
     2041 aacgtgctgc agatggcggg tgaatgcggc atcgtctacc tccggcattc ccagcacgta
     2101 aatcccgtcc agtccacaga ccagcgaaat cagccgccag gcgatatttt cggcgctatc
     2161 gcgcagggta aattcgccag cggcatggcc cgcgcgaatg atcctgaccg cttcgtcatg
     2221 ccacaggttc atggtcagca ggtacgcgct tttgatttcc ggatcgctgt cggccagcag
     2281 ctgcgcctgg cgccacaggc ggatataggg ctccagccgt ccgtcctcgc tgccgagggc
     2341 ggaaaacagc tgttcacgcc agccggcggt gcgcgaaagg cgctgcagat ccatcatctc
     2401 gcggatcacg cggataaacg cctgggattt tagctcgccg gaggaggtga aatggtggtg
     2461 gacctggcct gcggcgacgc ccgcggcggt ggcgatgttg cggaccgtca taccggtaaa
     2521 cccttgatcc agcgcgacgc gcatggcggc ctgcatgatg gtttcccgac gttcttcgcg
     2581 attcagatag cccatattgt cacctcctta tccacacatt atacgagccg gatgattaat
     2641 ggggaacctt accagagggc gccccagctg gcaattcctc ttaagtaagt aagagtatac
     2701 gtatatcggc taataacgta ttaaggcgct tcggcgcctt tttttatggg ggtattttca
     2761 tcccaatcca cacgtccaac gcacagcaaa caccacgtcg accctatcag ctgcgtgctt
     2821 tctatgagtc gttgctgcat aacaaaaaac tgcaaaaaat agtttgacag gacacgtgtc
     2881 caactttaag atgtacccaa agttggacac gtgtccaact ttgaattcaa aagatctttt
     2941 aagaaggaga tatactgatg atggaattta aagacttttc aacgggtctt tatgtagcag
     3001 ctaagttttc agaattaaca cttgatgcgc tggaagaact tcagcgctct ttacgtgttc
     3061 ctaatccagt tcctagagaa aaaattgcct cgactatatg ttattcaaga gtaaatgttc
     3121 catatgttcc atcgagtgga agttttgaag tagcttcttc tggacattta gaagtatgga
     3181 aaacacaaga tggatcgact cttgtacttg tgctagattc tgaatatctg cgctgtcgac
     3241 acatgtatgc gcgggcacta ggtgctacac atgattttga tgattacaca ccggcgataa
     3301 cattgtctta taatgttggg cccttatcat ttagcggtga tgtacaaatt ccggtcgtat
     3361 tagatcgtga atacaaagag cctcttaaac tcgattgggc agatgattta aaataaagga
     3421 tctccaggca tcaaataaaa cgaaaggctc agtcgaaaga ctgggccttt cgttttatct
     3481 gttgtttgtc ggtgaacgct ctctactaga gtcacactgg ctcaccttcg ggtgggcctt
     3541 tctgcgttta tagaagacca aggaggactt tggcaggtgc tctcctgagt aggacaaatc
     3601 cgccgcccta gacctagggt acgggttttg ctgcccgcaa acgggctgtt ctggtgttgc
     3661 tagtttgtta tcagaatcgc agatccggct tcagccggtt tgccggctga aagcactatt
     3721 tcttccagaa ttgccatgat tttttcccca cgggaggcgt cactggctcc cgtgttgtcg
     3781 gcagctttga ttcgataagc agcatcgcct gtttcaggct gtctatgtgt gactgttgag
     3841 ctgtaacaag ttgtctcagg tgttcaattt catgttctag ttgctttgtt ttactggttt
     3901 cacctgttct attaggtgtt acatgctgtt catctgttac attgtcgatc tgttcatggt
     3961 gaacagcttt gaatgcacca aaaactcgta aaagctctga tgtatctatc ttttttacac
     4021 cgttttcatc tgtgcatatg gacagttttc cctttgatat gtaacggtga acagttgttc
     4081 tacttttgtt tgttagtctt gatgcttcac tgatagatac aagagccata agaacctcag
     4141 atccttccgt atttagccag tatgttctct agtgtggttc gttgtttttg cgtgagccat
     4201 gagaacgaac cattgagatc atacttactt tgcatgtcac tcaaaaattt tgcctcaaaa
     4261 ctggtgagct gaatttttgc agttaaagca tcgtgtagtg tttttcttag tccgttatgt
     4321 aggtaggaat ctgatgtaat ggttgttggt attttgtcac cattcatttt tatctggttg
     4381 ttctcaagtt cggttacgag atccatttgt ctatctagtt caacttggaa aatcaacgta
     4441 tcagtcgggc ggcctcgctt atcaaccacc aatttcatat tgctgtaagt gtttaaatct
     4501 ttacttattg gtttcaaaac ccattggtta agccttttaa actcatggta gttattttca
     4561 agcattaaca tgaacttaaa ttcatcaagg ctaatctcta tatttgcctt gtgagttttc
     4621 ttttgtgtta gttcttttaa taaccactca taaatcctca tagagtattt gttttcaaaa
     4681 gacttaacat gttccagatt atattttatg aattttttta actggaaaag ataaggcaat
     4741 atctcttcac taaaaactaa ttctaatttt tcgcttgaga acttggcata gtttgtccac
     4801 tggaaaatct caaagccttt aaccaaagga ttcctgattt ccacagttct cgtcatcagc
     4861 tctctggttg ctttagctaa ta
//
