LOCUS       sequence-443193-        2643 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses gfp using J23119 promoter and pgRNA_AT backbone.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2643)
  AUTHORS   Sharma A, Omer Aden R, Puhar A, Cisneros DA
  TITLE     CRISPR-Cas-Guided Mutagenesis of Chromosome and Virulence Plasmid in
            Shigella flexneri by Cytosine Base Editing.
  JOURNAL   mSystems. 2022 Dec 21:e0104522. doi: 10.1128/msystems.01045-22.
  PUBMED    36541764
REFERENCE   2  (bases 1 to 2643)
  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: "mSystems. 
            2022 Dec 21:e0104522. doi: 10.1128/msystems.01045-22."
COMMENT     Sequence Label: pgRNA-R-gfp
FEATURES             Location/Qualifiers
     source          1..2643
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     184..203
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(350..373)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             complement(380..1093)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     RBS             1124..1135
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     promoter        complement(1155..1189)
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
     promoter        1202..1306
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1307..1375,1376..2167)
                     /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     complement(1525..1544)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(2338..2643,1..283)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
       61 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
      121 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
      181 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
      241 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
      301 ctttttacgg ttcctggcct tttgctggcc ttttgctcaa agcttaacct ttatcatcat
      361 cgtccttgta atcagatcct ttgtacagtt catccatacc atgcgtgatg cccgctgcgg
      421 ttacgaactc cagcagaacc atatgatcgc gtttctcgtt cggatcttta gacagaacgc
      481 tttgcgtgct cagatagtga ttgtctggca gcagaacagg accatcaccg attggagtgt
      541 tttgctggta gtgatcagcc agctgcacgc tgccatcctc cacgttgtgg cgaattttaa
      601 aattcgcttt aatgccattt ttttgtttat cggcggtgat gtaaacattg tggctgttaa
      661 aattgtattc cagcttatgg cccaggatat tgccgtcttc tttaaagtca atgcctttca
      721 gctcaatgcg gtttaccagg gtatcgcctt caaatttcac ttccgcacgc gttttgtacg
      781 tgccgtcatc cttaaaggaa atcgtgcgtt cctgcacata gccttccggc atggcggact
      841 tgaagaagtc atgctgcttc atatggtccg gataacgagc aaagcactga acaccataag
      901 tcagcgtcgt taccagagtc ggccaaggaa ccggcagttt accagtagta cagatgaact
      961 tcagcgtcag tttaccatta gttgcgtcac cttcaccctc gccacgcacg gaaaacttat
     1021 gaccgttgac atcaccatcc agttccacca gaatagggac gacaccagtg aacagctctt
     1081 cgcctttacg cattatttta aatttttttg aatccatcta gtatttctcc tctttcggat
     1141 ccacgcgtgg tctcactagt attataccta ggactgagct agctgtcaag aattcaatgt
     1201 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
     1261 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
     1321 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
     1381 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
     1441 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
     1501 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
     1561 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
     1621 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
     1681 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     1741 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     1801 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     1861 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     1921 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     1981 tggctggttt attgctgata aatctggagc cggtgagcgt gggtcgcgcg gtatcattgc
     2041 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     2101 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     2161 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     2221 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     2281 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     2341 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     2401 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     2461 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     2521 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     2581 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     2641 gca
//
