LOCUS       sequence-490797-        5102 bp    DNA     circular SYN 27-OCT-2025
DEFINITION  Bacterial expression of RFP under control of the pJEX promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5102)
  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 5102)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 27, 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: pBA1326
FEATURES             Location/Qualifiers
     source          1..5102
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      253..476
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             complement(1019..1834)
                     /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(1019..1834)
                     /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     1742..1761
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1826..1845
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     RBS             2603..2611
                     /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)"
     CDS             2959..3636
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     terminator      3652..3723
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3739..3766
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(join(4357..5102,1..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 gttttttttt gtttatattc aagtggttat aatttataga
      421 ataaagaaag aataaaaaaa gataaaaaga atagatccca gccctgtgta taactcacta
      481 ctttagtcag ttccgcagta ttacaaaagg atgtcgcaaa cgctgtttgc tcctctacaa
      541 aacagacctt aaaaccctaa aggcttaagt agcaccctcg caagctcggg caaatcgctg
      601 aatattcctt ttgtctccga ccatcaggca cctgagtcgc tgtctttttc gtgacattca
      661 gttcgctgcg ctcacggctc tggcagtgaa tgggggtaaa tggcactaca ggcgcctttt
      721 atggattcat gcaaggaaac tacccataat acaagaaaag cccgtcacgg gcttctcagg
      781 gcgttttatg gcgggtctgc tatgtggtgc tatctgactt tttgctgttc agcagttcct
      841 gccctctgat tttccagtct gaccacttcg gattatcccg tgacaggtca ttcagactgg
      901 ctaatgcacc cagtaaggca gcggtatcat caacaggctt acccgtctta ctgtccctgt
      961 gtcagatttc ttactttatt caggcgatta cgtgttacaa ccaattaacc aattctgatt
     1021 agaaaaactc atcgagcatc aaatgaaact gcaatttatt catatcagga ttatcaatac
     1081 catatttttg aaaaagccgt ttctgtaatg aaggagaaaa ctcaccgagg cagttccata
     1141 ggatggcaag atcctggtat cggtctgcga ttccgactcg tccaacatca atacaaccta
     1201 ttaatttccc ctcgtcaaaa ataaggttat caagtgagaa atcaccatga gtgacgactg
     1261 aatccggtga gaatggcaaa agcttatgca tttctttcca gacttgttca acaggccagc
     1321 cattacgctc gtcatcaaaa tcactcgcat caaccaaacc gttattcatt cgtgattgcg
     1381 cctgagcgag gcgaaatacg cgatcgctgt taaaaggaca attacaaaca ggaatcgaat
     1441 gcaaccggcg caggaacact gccagcgcat caacaatatt ttcacctgaa tcaggatatt
     1501 cttctaatac ctggaatgct gttttcccgg ggatcgcagt ggtgagtaac catgcatcat
     1561 caggagtacg gataaaatgc ttgatggtcg gaagaggcat aaattccgtc agccagttta
     1621 gtctgaccat ctcatctgta acatcattgg caacgctacc tttgccatgt ttcagaaaca
     1681 actctggcgc atcgggcttc ccatacaatc gatagattgt cgcacctgat tgcccgacat
     1741 tatcgcgagc ccatttatac ccatataaat cagcatccat gttggaattt aatcgcggcc
     1801 tggagcaaga cgtttcccgt tgaatatggc tcataacacc ccttgtatta ctgtttatgt
     1861 aagcagacag ttttattgtt catgatgata tatttttatc ttgtgcaatg taacatcaga
     1921 gattttgaga cacaacgtgg ctttgttgaa taaatcgaac ttttgctgag ttgaaggatc
     1981 agcacctgcc cgaaccagag acttatctca ggacgtctta cgaaaataac tcaagctgaa
     2041 taacgtgctg cagatggcgg gtgaatgcgg catcgtctac ctccggcatt cccagcacgt
     2101 aaatcccgtc cagtccacag accagcgaaa tcagccgcca ggcgatattt tcggcgctat
     2161 cgcgcagggt aaattcgcca gcggcatggc ccgcgcgaat gatcctgacc gcttcgtcat
     2221 gccacaggtt catggtcagc aggtacgcgc ttttgatttc cggatcgctg tcggccagca
     2281 gctgcgcctg gcgccacagg cggatatagg gctccagccg tccgtcctcg ctgccgaggg
     2341 cggaaaacag ctgttcacgc cagccggcgg tgcgcgaaag gcgctgcaga tccatcatct
     2401 cgcggatcac gcggataaac gcctgggatt ttagctcgcc ggaggaggtg aaatggtggt
     2461 ggacctggcc tgcggcgacg cccgcggcgg tggcgatgtt gcggaccgtc ataccggtaa
     2521 acccttgatc cagcgcgacg cgcatggcgg cctgcatgat ggtttcccga cgttcttcgc
     2581 gattcagata gcccatattg tcacctcctt atccacacat tatacgagcc ggatgattaa
     2641 tggggaacct taccagaggg cgccccagct ggcaattcct cttaagtaag taagagtata
     2701 cgtatatcgg ctaataacgt attaaggcgc ttcggcgcct ttttttatgg gggtattttc
     2761 atcccaatcc acacgtccaa cgcacagcaa acaccacgtc gaccctatca gctgcgtgct
     2821 ttctatgagt cgttgctgca taacaaaaaa ctgcaaaaaa tagtttgaca ggacacgtgt
     2881 ccaactttaa gatgtaccca aagttggaca cgtgtccaac tttgaattca aaagatcttt
     2941 taagaaggag atattctgat ggcgagtagc gaagacgtta tcaaagagtt catgcgtttc
     3001 aaagttcgta tggaaggttc cgttaacggt cacgagttcg aaatcgaagg tgaaggtgaa
     3061 ggtcgtccgt acgaaggtac ccagaccgct aaactgaaag ttaccaaagg tggtccgctg
     3121 ccgttcgctt gggacatcct gtccccgcag ttccagtacg gttccaaagc ttacgttaaa
     3181 cacccggctg acatcccgga ctacctgaaa ctgtccttcc cggaaggttt caaatgggaa
     3241 cgtgttatga acttcgaaga cggtggtgtt gttaccgtta cccaggactc ctccctgcaa
     3301 gacggtgagt tcatctacaa agttaaactg cgtggtacca acttcccgtc cgacggtccg
     3361 gttatgcaga aaaaaaccat gggttgggaa gcttccaccg aacgtatgta cccggaagac
     3421 ggtgctctga aaggtgaaat caaaatgcgt ctgaaactga aagacggtgg tcactacgac
     3481 gctgaagtta aaaccaccta catggctaaa aaaccggttc agctgccggg tgcttacaaa
     3541 accgacatca aactggacat cacctcccac aacgaagact acaccatcgt tgaacagtac
     3601 gaacgtgctg aaggtcgtca ctccaccggt gcttaaagga tctccaggca tcaaataaaa
     3661 cgaaaggctc agtcgaaaga ctgggccttt cgttttatct gttgtttgtc ggtgaacgct
     3721 ctctactaga gtcacactgg ctcaccttcg ggtgggcctt tctgcgttta tagaagacca
     3781 aggaggactt tggcaggtgc tctcctgagt aggacaaatc cgccgcccta gacctagggt
     3841 acgggttttg ctgcccgcaa acgggctgtt ctggtgttgc tagtttgtta tcagaatcgc
     3901 agatccggct tcagccggtt tgccggctga aagcactatt tcttccagaa ttgccatgat
     3961 tttttcccca cgggaggcgt cactggctcc cgtgttgtcg gcagctttga ttcgataagc
     4021 agcatcgcct gtttcaggct gtctatgtgt gactgttgag ctgtaacaag ttgtctcagg
     4081 tgttcaattt catgttctag ttgctttgtt ttactggttt cacctgttct attaggtgtt
     4141 acatgctgtt catctgttac attgtcgatc tgttcatggt gaacagcttt gaatgcacca
     4201 aaaactcgta aaagctctga tgtatctatc ttttttacac cgttttcatc tgtgcatatg
     4261 gacagttttc cctttgatat gtaacggtga acagttgttc tacttttgtt tgttagtctt
     4321 gatgcttcac tgatagatac aagagccata agaacctcag atccttccgt atttagccag
     4381 tatgttctct agtgtggttc gttgtttttg cgtgagccat gagaacgaac cattgagatc
     4441 atacttactt tgcatgtcac tcaaaaattt tgcctcaaaa ctggtgagct gaatttttgc
     4501 agttaaagca tcgtgtagtg tttttcttag tccgttatgt aggtaggaat ctgatgtaat
     4561 ggttgttggt attttgtcac cattcatttt tatctggttg ttctcaagtt cggttacgag
     4621 atccatttgt ctatctagtt caacttggaa aatcaacgta tcagtcgggc ggcctcgctt
     4681 atcaaccacc aatttcatat tgctgtaagt gtttaaatct ttacttattg gtttcaaaac
     4741 ccattggtta agccttttaa actcatggta gttattttca agcattaaca tgaacttaaa
     4801 ttcatcaagg ctaatctcta tatttgcctt gtgagttttc ttttgtgtta gttcttttaa
     4861 taaccactca taaatcctca tagagtattt gttttcaaaa gacttaacat gttccagatt
     4921 atattttatg aattttttta actggaaaag ataaggcaat atctcttcac taaaaactaa
     4981 ttctaatttt tcgcttgaga acttggcata gtttgtccac tggaaaatct caaagccttt
     5041 aaccaaagga ttcctgattt ccacagttct cgtcatcagc tctctggttg ctttagctaa
     5101 ta
//
