LOCUS       sequence-353896-        3118 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  sgRNA targeting RFP for repression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3118)
  AUTHORS   Tickman BI, Burbano DA, Chavali VP, Kiattisewee C, Fontana J, 
            Khakimzhan A, Noireaux V, Zalatan JG, Carothers JM
  TITLE     Multi-layer CRISPRa/i circuits for dynamic genetic programs in 
            cell-free and bacterial systems.
  JOURNAL   Cell Syst. 2021 Nov 17. pii: S2405-4712(21)00419-1. doi: 
            10.1016/j.cels.2021.10.008.
  PUBMED    34800362
REFERENCE   2  (bases 1 to 3118)
  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: "Cell Syst. 
            2021 Nov 17. pii: S2405-4712(21)00419-1. doi: 
            10.1016/j.cels.2021.10.008."
COMMENT     Sequence Label: pBT009.J3.RR2
FEATURES             Location/Qualifiers
     source          1..3118
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(449..1240,1241..1309))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     1072..1091
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1310..1414)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1482..1500
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2022..2056
                     /label=BBa_J23117
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.06"
     misc_RNA        2077..2152
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(2637..2654)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(2808..3118,1..278))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2888..2907)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
       61 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
      121 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
      181 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
      241 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
      301 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
      361 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
      421 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
      481 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
      541 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
      601 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
      661 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
      721 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
      781 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
      841 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
      901 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
      961 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     1021 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaacacgg gataataccg
     1081 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     1141 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     1201 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     1261 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     1321 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     1381 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     1441 acgtctaaga aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc
     1501 cctttcgtct tcaagaattc tggcgaatcc tctgaccagc cagaaaacga cctttctgtg
     1561 gtgaaaccgg atgctgcaat tcagagcgcc agcaagtggg ggacagcaga agacctgacc
     1621 gccgcagagt ggatgtttga catggtgaag actatcgcac catcagccag aaaaccgaat
     1681 tttgctgggt gggctaacga tatccgcctg atgcgtgaac gtgacggacg taaccaccgc
     1741 gacatgtgtg tgctgttccg ctgggcatgc caggacaact tctggtccgg taacgtgctg
     1801 agcccggcca agcttagccc gaaaagtgcc acctgtggca attccgacgt cagcatttgc
     1861 gatcattcac gcagcgctta ttcagttgct cactgcgatg tcataatcat cgctacgagc
     1921 tgtgaaagat gcataaagct cgtacgacgc gttcgctcgt ctcctcactt ctcctacgga
     1981 gcgttctgga cacaacgtcg tcttgaagtt gcgattatag attgacagct agctcagtcc
     2041 tagggattgt gctagctgga accgtactgg aactgcgttt tagagctaga aatagcaagt
     2101 taaaataagg ctagtccgtt atcaacttga aaaagtggca ccgagtcggt gcttttttta
     2161 acgcatgaga aagcccccgg aagatcacct tccgggggct tttttattgc gcggatcctt
     2221 actccgtcga caagcttgcg gccgcactcg aggaattccg actcaattag ttcagtcagt
     2281 ttcaggatat tagtcatctc tacattgatt atgagtattc agaaattcct taaatattct
     2341 gacaaatgct ctttccctaa actcccccca taaaaaaacc cgccgaagcg ggtttttacg
     2401 ttatttgcgg attaacgatt actcgttatc agaaccgccc agacctgcgt tcagcagttc
     2461 tgccaggctg gcagatgcgt cttccggaat tgatccgtcg accgatgccc ttgagagcct
     2521 tcaacccagt cagctccttc cggtgggcgc ggggcatgac tatcgtcgcc gcacttatga
     2581 ctgtcttctt tatcatgcaa ctcgtaggac aggtgccggc agcgctcttc cgcttcctcg
     2641 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     2701 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     2761 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     2821 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     2881 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     2941 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     3001 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     3061 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttg
//
