LOCUS       sequence-488909-        4690 bp    DNA     circular SYN 09-OCT-2025
DEFINITION  Bacterial expression of mutant T4 Acb2 (Y8A) under control of the 
            pJex promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4690)
  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 4690)
  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: pBA1543
FEATURES             Location/Qualifiers
     source          1..4690
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      397..468
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      484..511
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(1102..2052)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      2100..2322
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             complement(2865..3680)
                     /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(2865..3680)
                     /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     3588..3607
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3672..3691
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     RBS             4449..4457
                     /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)"
ORIGIN
        1 aaaaaactgc aaaaaatagt ttgacaggac acgtgtccaa ctttaagatg tacccaaagt
       61 tggacacgtg tccaactttg aattcaaaag atcttttaag aaggagatat actgatgatt
      121 gaagatatta agggtgcgaa accacatact gaagagaaaa tcggtaaagt aaatgctatt
      181 aaagacgctg aagttcgttt aggacttatc tttgatgctt tatatgatga attctgggaa
      241 gcactagata attgcgaaga ctgtgaattc gcgaagaatt atgctgaaag tctcgatcag
      301 ttaactattg ctaaaacgaa actcaaagaa gccagtatgt gggcttgtcg tgcagtgttc
      361 caaccagagg aaaaatacta aaggatctcc aggcatcaaa taaaacgaaa ggctcagtcg
      421 aaagactggg cctttcgttt tatctgttgt ttgtcggtga acgctctcta ctagagtcac
      481 actggctcac cttcgggtgg gcctttctgc gtttatagaa gaccaaggag gactttggca
      541 ggtgctctcc tgagtaggac aaatccgccg ccctagacct agggtacggg ttttgctgcc
      601 cgcaaacggg ctgttctggt gttgctagtt tgttatcaga atcgcagatc cggcttcagc
      661 cggtttgccg gctgaaagca ctatttcttc cagaattgcc atgatttttt ccccacggga
      721 ggcgtcactg gctcccgtgt tgtcggcagc tttgattcga taagcagcat cgcctgtttc
      781 aggctgtcta tgtgtgactg ttgagctgta acaagttgtc tcaggtgttc aatttcatgt
      841 tctagttgct ttgttttact ggtttcacct gttctattag gtgttacatg ctgttcatct
      901 gttacattgt cgatctgttc atggtgaaca gctttgaatg caccaaaaac tcgtaaaagc
      961 tctgatgtat ctatcttttt tacaccgttt tcatctgtgc atatggacag ttttcccttt
     1021 gatatgtaac ggtgaacagt tgttctactt ttgtttgtta gtcttgatgc ttcactgata
     1081 gatacaagag ccataagaac ctcagatcct tccgtattta gccagtatgt tctctagtgt
     1141 ggttcgttgt ttttgcgtga gccatgagaa cgaaccattg agatcatact tactttgcat
     1201 gtcactcaaa aattttgcct caaaactggt gagctgaatt tttgcagtta aagcatcgtg
     1261 tagtgttttt cttagtccgt tatgtaggta ggaatctgat gtaatggttg ttggtatttt
     1321 gtcaccattc atttttatct ggttgttctc aagttcggtt acgagatcca tttgtctatc
     1381 tagttcaact tggaaaatca acgtatcagt cgggcggcct cgcttatcaa ccaccaattt
     1441 catattgctg taagtgttta aatctttact tattggtttc aaaacccatt ggttaagcct
     1501 tttaaactca tggtagttat tttcaagcat taacatgaac ttaaattcat caaggctaat
     1561 ctctatattt gccttgtgag ttttcttttg tgttagttct tttaataacc actcataaat
     1621 cctcatagag tatttgtttt caaaagactt aacatgttcc agattatatt ttatgaattt
     1681 ttttaactgg aaaagataag gcaatatctc ttcactaaaa actaattcta atttttcgct
     1741 tgagaacttg gcatagtttg tccactggaa aatctcaaag cctttaacca aaggattcct
     1801 gatttccaca gttctcgtca tcagctctct ggttgcttta gctaatacac cataagcatt
     1861 ttccctactg atgttcatca tctgagcgta ttggttataa gtgaacgata ccgtccgttc
     1921 tttccttgta gggttttcaa tcgtggggtt gagtagtgcc acacagcata aaattagctt
     1981 ggtttcatgc tccgttaagt catagcgact aatcgctagt tcatttgctt tgaaaacaac
     2041 taattcagac atacatctca attggtctag gtgattttaa tcactatacc aattgagatg
     2101 ggctagtcaa tgataattac tagtcctttt cctttgagtt gtgggtatct gtaaattctg
     2161 ctagaccttt gctggaaaac ttgtaaattc tgctataccc tctgtaaatt ccgctagacc
     2221 tttgtgtgtt ttttttgttt atattcaagt ggttataatt tatagaataa agaaagaata
     2281 aaaaaagata aaaagaatag atcccagccc tgtgtataac tcactacttt agtcagttcc
     2341 gcagtattac aaaaggatgt cgcaaacgct gtttgctcct ctacaaaaca gaccttaaaa
     2401 ccctaaaggc ttaagtagca ccctcgcaag ctcgggcaaa tcgctgaata ttccttttgt
     2461 ctccgaccat caggcacctg agtcgctgtc tttttcgtga cattcagttc gctgcgctca
     2521 cggctctggc agtgaatggg ggtaaatggc actacaggcg ccttttatgg attcatgcaa
     2581 ggaaactacc cataatacaa gaaaagcccg tcacgggctt ctcagggcgt tttatggcgg
     2641 gtctgctatg tggtgctatc tgactttttg ctgttcagca gttcctgccc tctgattttc
     2701 cagtctgacc acttcggatt atcccgtgac aggtcattca gactggctaa tgcacccagt
     2761 aaggcagcgg tatcatcaac aggcttaccc gtcttactgt ccctgtgtca gatttcttac
     2821 tttattcagg cgattacgtg ttacaaccaa ttaaccaatt ctgattagaa aaactcatcg
     2881 agcatcaaat gaaactgcaa tttattcata tcaggattat caataccata tttttgaaaa
     2941 agccgtttct gtaatgaagg agaaaactca ccgaggcagt tccataggat ggcaagatcc
     3001 tggtatcggt ctgcgattcc gactcgtcca acatcaatac aacctattaa tttcccctcg
     3061 tcaaaaataa ggttatcaag tgagaaatca ccatgagtga cgactgaatc cggtgagaat
     3121 ggcaaaagct tatgcatttc tttccagact tgttcaacag gccagccatt acgctcgtca
     3181 tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg attgcgcctg agcgaggcga
     3241 aatacgcgat cgctgttaaa aggacaatta caaacaggaa tcgaatgcaa ccggcgcagg
     3301 aacactgcca gcgcatcaac aatattttca cctgaatcag gatattcttc taatacctgg
     3361 aatgctgttt tcccggggat cgcagtggtg agtaaccatg catcatcagg agtacggata
     3421 aaatgcttga tggtcggaag aggcataaat tccgtcagcc agtttagtct gaccatctca
     3481 tctgtaacat cattggcaac gctacctttg ccatgtttca gaaacaactc tggcgcatcg
     3541 ggcttcccat acaatcgata gattgtcgca cctgattgcc cgacattatc gcgagcccat
     3601 ttatacccat ataaatcagc atccatgttg gaatttaatc gcggcctgga gcaagacgtt
     3661 tcccgttgaa tatggctcat aacacccctt gtattactgt ttatgtaagc agacagtttt
     3721 attgttcatg atgatatatt tttatcttgt gcaatgtaac atcagagatt ttgagacaca
     3781 acgtggcttt gttgaataaa tcgaactttt gctgagttga aggatcagca cctgcccgaa
     3841 ccagagactt atctcaggac gtcttacgaa aataactcaa gctgaataac gtgctgcaga
     3901 tggcgggtga atgcggcatc gtctacctcc ggcattccca gcacgtaaat cccgtccagt
     3961 ccacagacca gcgaaatcag ccgccaggcg atattttcgg cgctatcgcg cagggtaaat
     4021 tcgccagcgg catggcccgc gcgaatgatc ctgaccgctt cgtcatgcca caggttcatg
     4081 gtcagcaggt acgcgctttt gatttccgga tcgctgtcgg ccagcagctg cgcctggcgc
     4141 cacaggcgga tatagggctc cagccgtccg tcctcgctgc cgagggcgga aaacagctgt
     4201 tcacgccagc cggcggtgcg cgaaaggcgc tgcagatcca tcatctcgcg gatcacgcgg
     4261 ataaacgcct gggattttag ctcgccggag gaggtgaaat ggtggtggac ctggcctgcg
     4321 gcgacgcccg cggcggtggc gatgttgcgg accgtcatac cggtaaaccc ttgatccagc
     4381 gcgacgcgca tggcggcctg catgatggtt tcccgacgtt cttcgcgatt cagatagccc
     4441 atattgtcac ctccttatcc acacattata cgagccggat gattaatggg gaaccttacc
     4501 agagggcgcc ccagctggca attcctctta agtaagtaag agtatacgta tatcggctaa
     4561 taacgtatta aggcgcttcg gcgccttttt ttatgggggt attttcatcc caatccacac
     4621 gtccaacgca cagcaaacac cacgtcgacc ctatcagctg cgtgctttct atgagtcgtt
     4681 gctgcataac
//
