LOCUS       sequence-487439-        4556 bp    DNA     circular SYN 29-SEP-2025
DEFINITION  Reporter J1-J3(122)-BBa_J23117 with Sth1 PAM.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4556)
  AUTHORS   Kiattisewee C, Karanjia AV, Cardiff RAL, Olander KE, Leejareon P, 
            Alvi SS, Carothers JM, Zalatan JG
  TITLE     Systematic Mapping of Bacterial CRISPRa Systems for Synergistic Gene
            Activation Reveals Antagonistic Effects.
  JOURNAL   ACS Synth Biol. 2025 Jul 22. doi: 10.1021/acssynbio.5c00358.
  PUBMED    40693287
REFERENCE   2  (bases 1 to 4556)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 29, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "ACS Synth 
            Biol. 2025 Jul 22. doi: 10.1021/acssynbio.5c00358."
COMMENT     Sequence Label: pKO023
FEATURES             Location/Qualifiers
     source          1..4556
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        437..471
                     /label=BBa_J23117
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.06"
     RBS             481..492
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             499..1176
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     terminator      1209..1280
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1296..1323
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(1854..2804)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     terminator      complement(3566..3660)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(join(3690..4481,4482..4550))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4313..4332
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(4551..4556,1..99))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
       61 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
      121 acctgtggca attccgacgt cgcctacggt atccaccgga gacctatggc agcctccggc
      181 cgccatagga cacctttggt tgccaagggt gacctatggt gaccatgggc caccacgggc
      241 gacctcaggt atcctgcggt gtcctgcggt taccaaaggc gtcctttggg ttccaccgga
      301 tacctccgga cgtcataatc atcgctacga gctgtgaaag atgcataaag ctcgtacgac
      361 gcgttcgctc gtctcctcac ttcattctca acggagcgtt ctggacacaa cgtcgtcttg
      421 aagttgcgat tatagattga cagctagctc agtcctaggg attgtgctag cgaattcatt
      481 aaagaggaga aaggtaccat ggcgagtagc gaagacgtta tcaaagagtt catgcgtttc
      541 aaagttcgta tggaaggttc cgttaacggt cacgagttcg aaatcgaagg tgaaggtgaa
      601 ggtcgtccgt acgaaggtac ccagaccgct aaactgaaag ttaccaaagg tggtccgctg
      661 ccgttcgctt gggacatcct gtccccgcag ttccagtacg gttccaaagc ttacgttaaa
      721 cacccggctg acatcccgga ctacctgaaa ctgtccttcc cggaaggttt caaatgggaa
      781 cgtgttatga acttcgaaga cggtggtgtt gttaccgtta cccaggactc ctccctgcaa
      841 gacggtgagt tcatctacaa agttaaactg cgtggtacca acttcccgtc cgacggtccg
      901 gttatgcaga aaaaaaccat gggttgggaa gcttccaccg aacgtatgta cccggaagac
      961 ggtgctctga aaggtgaaat caaaatgcgt ctgaaactga aagacggtgg tcactacgac
     1021 gctgaagtta aaaccaccta catggctaaa aaaccggttc agctgccggg tgcttacaaa
     1081 accgacatca aactggacat cacctcccac aacgaagact acaccatcgt tgaacagtac
     1141 gaacgtgctg aaggtcgtca ctccaccggt gcttaaggat ccaaactcga gtaaggatct
     1201 ccaggcatca aataaaacga aaggctcagt cgaaagactg ggcctttcgt tttatctgtt
     1261 gtttgtcggt gaacgctctc tactagagtc acactggctc accttcgggt gggcctttct
     1321 gcgtttatac ctagggtacg ggttttgctg cccgcaaacg ggctgttctg gtgttgctag
     1381 tttgttatca gaatcgcaga tccggcttca gccggtttgc cggctgaaag cgctatttct
     1441 tccagaattg ccatgatttt ttccccacgg gaggcgtcac tggctcccgt gttgtcggca
     1501 gctttgattc gataagcagc atcgcctgtt tcaggctgtc tatgtgtgac tgttgagctg
     1561 taacaagttg tctcaggtgt tcaatttcat gttctagttg ctttgtttta ctggtttcac
     1621 ctgttctatt aggtgttaca tgctgttcat ctgttacatt gtcgatctgt tcatggtgaa
     1681 cagctttgaa tgcaccaaaa actcgtaaaa gctctgatgt atctatcttt tttacaccgt
     1741 tttcatctgt gcatatggac agttttccct ttgatatgta acggtgaaca gttgttctac
     1801 ttttgtttgt tagtcttgat gcttcactga tagatacaag agccataaga acctcagatc
     1861 cttccgtatt tagccagtat gttctctagt gtggttcgtt gtttttgcgt gagccatgag
     1921 aacgaaccat tgagatcata cttactttgc atgtcactca aaaattttgc ctcaaaactg
     1981 gtgagctgaa tttttgcagt taaagcatcg tgtagtgttt ttcttagtcc gttatgtagg
     2041 taggaatctg atgtaatggt tgttggtatt ttgtcaccat tcatttttat ctggttgttc
     2101 tcaagttcgg ttacgagatc catttgtcta tctagttcaa cttggaaaat caacgtatca
     2161 gtcgggcggc ctcgcttatc aaccaccaat ttcatattgc tgtaagtgtt taaatcttta
     2221 cttattggtt tcaaaaccca ttggttaagc cttttaaact catggtagtt attttcaagc
     2281 attaacatga acttaaattc atcaaggcta atctctatat ttgccttgtg agttttcttt
     2341 tgtgttagtt cttttaataa ccactcataa atcctcatag agtatttgtt ttcaaaagac
     2401 ttaacatgtt ccagattata ttttatgaat ttttttaact ggaaaagata aggcaatatc
     2461 tcttcactaa aaactaattc taatttttcg cttgagaact tggcatagtt tgtccactgg
     2521 aaaatctcaa agcctttaac caaaggattc ctgatttcca cagttctcgt catcagctct
     2581 ctggttgctt tagctaatac accataagca ttttccctac tgatgttcat catctgagcg
     2641 tattggttat aagtgaacga taccgtccgt tctttccttg tagggttttc aatcgtgggg
     2701 ttgagtagtg ccacacagca taaaattagc ttggtttcat gctccgttaa gtcatagcga
     2761 ctaatcgcta gttcatttgc tttgaaaaca actaattcag acatacatct caattggtct
     2821 aggtgatttt aatcactata ccaattgaga tgggctagtc aatgataatt actagtcctt
     2881 ttcccgggtg atctgggtat ctgtaaattc tgctagacct ttgctggaaa acttgtaaat
     2941 tctgctagac cctctgtaaa ttccgctaga cctttgtgtg ttttttttgt ttatattcaa
     3001 gtggttataa tttatagaat aaagaaagaa taaaaaaaga taaaaagaat agatcccagc
     3061 cctgtgtata actcactact ttagtcagtt ccgcagtatt acaaaaggat gtcgcaaacg
     3121 ctgtttgctc ctctacaaaa cagaccttaa aaccctaaag gcttaagtag caccctcgca
     3181 agctcgggca aatcgctgaa tattcctttt gtctccgacc atcaggcacc tgagtcgctg
     3241 tctttttcgt gacattcagt tcgctgcgct cacggctctg gcagtgaatg ggggtaaatg
     3301 gcactacagg cgccttttat ggattcatgc aaggaaacta cccataatac aagaaaagcc
     3361 cgtcacgggc ttctcagggc gttttatggc gggtctgcta tgtggtgcta tctgactttt
     3421 tgctgttcag cagttcctgc cctctgattt tccagtctga ccacttcgga ttatcccgtg
     3481 acaggtcatt cagactggct aatgcaccca gtaaggcagc ggtatcatca acaggcttac
     3541 ccgtcttact gtccctagtg cttggattct caccaataaa aaacgcccgg cggcaaccga
     3601 gcgttctgaa caaatccaga tggagttctg aggtcattac tggatctatc aacaggagtc
     3661 caagcgagct cgtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     3721 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     3781 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     3841 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     3901 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     3961 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     4021 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     4081 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     4141 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     4201 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     4261 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     4321 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     4381 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     4441 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     4501 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actctt
//
