LOCUS       sequence-403410-        3367 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Low-copy constitutively expressed sfGFP for E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3367)
  TITLE     Transcriptional Fluoride Riboswitch for E. coli Expression
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 3367)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pJBL811
FEATURES             Location/Qualifiers
     source          1..3367
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      96..190
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     primer_bind     complement(597..614)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(731..1276)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(811..830)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        1470..1504
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
     CDS             join(1560..1562,1563..2276)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             1560..2273
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Nager et al., 2011)"
                     /label=superfolder GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(1590..1618)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     2213..2233
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     CDS             2296..2325
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2341..2358
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2414..2431)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      2584..2630
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             join(2783..3367,1..75)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2849..2868)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
ORIGIN
        1 tgtgatggct tccatgtcgg cagaatgctt aatgaattac aacagtactg cgatgagtgg
       61 cagggcgggg cgtaatttga tatcgagctc gcttggactc ctgttgatag atccagtaat
      121 gacctcagaa ctccatctgg atttgttcag aacgctcggt tgccgccggg cgttttttat
      181 tggtgagaat ccaagcctcc gatcaacgtc tcattttcgc caaaagttgg cccagggctt
      241 cccggtatca acagggacac caggatttat ttattctgcg aagtgatctt ccgtcacagg
      301 tatttattcg gcgcaaagtg cgtcgggtga tgctgccaac ttactgattt agtgtatgat
      361 ggtgtttttg aggtgctcca gtggcttctg tttctatcag ctgtccctcc tgttcagcta
      421 ctgacggggt ggtgcgtaac ggcaaaagca ccgccggaca tcagcgctag cggagtgtat
      481 actggcttac tatgttggca ctgatgaggg tgtcagtgaa gtgcttcatg tggcaggaga
      541 aaaaaggctg caccggtgcg tcagcagaat atgtgataca ggatatattc cgcttcctcg
      601 ctcactgact cgctacgctc ggtcgttcga ctgcggcgag cggaaatggc ttacgaacgg
      661 ggcggagatt tcctggaaga tgccaggaag atacttaaca gggaagtgag agggccgcgg
      721 caaagccgtt tttccatagg ctccgccccc ctgacaagca tcacgaaatc tgacgctcaa
      781 atcagtggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggcggct
      841 ccctcgtgcg ctctcctgtt cctgcctttc ggtttaccgg tgtcattccg ctgttatggc
      901 cgcgtttgtc tcattccacg cctgacactc agttccgggt aggcagttcg ctccaagctg
      961 gactgtatgc acgaaccccc cgttcagtcc gaccgctgcg ccttatccgg taactatcgt
     1021 cttgagtcca acccggaaag acatgcaaaa gcaccactgg cagcagccac tggtaattga
     1081 tttagaggag ttagtcttga agtcatgcgc cggttaaggc taaactgaaa ggacaagttt
     1141 tggtgactgc gctcctccaa gccagttacc tcggttcaaa gagttggtag ctcagagaac
     1201 cttcgaaaaa ccgccctgca aggcggtttt ttcgttttca gagcaagaga ttacgcgcag
     1261 accaaaacga tctcaagaag atcatcttat taatcagata aaatatttct agatttcagt
     1321 gcaatttatc tcttcaaatg tagcacctga agtcagcccc atacgatata agttgtaatt
     1381 ctcatgtttg acagcttatc atcgataagc ttccgatggc gcgccgagag gctttacact
     1441 ttatgcttcc ggctgaattc taaagatctt tgacagctag ctcagtccta ggtataatac
     1501 tagtacgtcg actctcgagt gagattgttg acggtaccgt attttaggag gaaggatcta
     1561 tgagcaaagg agaagaactt ttcactggag ttgtcccaat tcttgttgaa ttagatggtg
     1621 atgttaatgg gcacaaattt tctgtccgtg gagagggtga aggtgatgct acaaacggaa
     1681 aactcaccct taaatttatt tgcactactg gaaaactacc tgttccgtgg ccaacacttg
     1741 tcactactct gacctatggt gttcaatgct tttcccgtta tccggatcac atgaaacggc
     1801 atgacttttt caagagtgcc atgcccgaag gttatgtaca ggaacgcact atatctttca
     1861 aagatgacgg gacctacaag acgcgtgctg aagtcaagtt tgaaggtgat acccttgtta
     1921 atcgtatcga gttaaagggt attgatttta aagaagatgg aaacattctt ggacacaaac
     1981 tcgagtacaa ctttaactca cacaatgtat acatcacggc agacaaacaa aagaatggaa
     2041 tcaaagctaa cttcaaaatt cgccacaacg ttgaagatgg ttccgttcaa ctagcagacc
     2101 attatcaaca aaatactcca attggcgatg gccctgtcct tttaccagac aaccattacc
     2161 tgtcgacaca atctgtcctt tcgaaagatc ccaacgaaaa gcgtgaccac atggtccttc
     2221 ttgagtttgt aactgctgct gggattacac atggcatgga tgagctctac aaataaggat
     2281 ctgaagcttg ggcccgaaca aaaactcatc tcagaagagg atctgaatag cgccgtcgac
     2341 catcatcatc atcatcattg agtttaaacg gtctccagct tggctgtttt ggcggatgag
     2401 agaagatttt cagcctgata cagattaaat cagaacgcag aagcggtctg ataaaacaga
     2461 atttgcctgg cggcagtagc gcggtggtcc cacctgaccc catgccgaac tcagaagtga
     2521 aacgccgtag cgccgatggt agtgtggggt ctccccatgc gagagtaggg aactgccagg
     2581 catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg
     2641 tcggtgaact ggatccttac tcgagtctag actgcagttg atcgggcacg taagaggttc
     2701 caactttcac cataatgaaa taagatcact accgggcgta ttttttgagt tatcgagatt
     2761 ttcaggagct aaggaagcta aaatggagaa aaaaatcact ggatatacca ccgttgatat
     2821 atcccaatgg catcgtaaag aacattttga ggcatttcag tcagttgctc aatgtaccta
     2881 taaccagacc gttcagctgg atattacggc ctttttaaag accgtaaaga aaaataagca
     2941 caagttttat ccggccttta ttcacattct tgcccgcctg atgaatgctc atccggaatt
     3001 tcgtatggca atgaaagacg gtgagctggt gatatgggat agtgttcacc cttgttacac
     3061 cgttttccat gagcaaactg aaacgttttc atcgctctgg agtgaatacc acgacgattt
     3121 ccggcagttt ctacacatat attcgcaaga tgtggcgtgt tacggtgaaa acctggccta
     3181 tttccctaaa gggtttattg agaatatgtt tttcgtctca gccaatccct gggtgagttt
     3241 caccagtttt gatttaaacg tggccaatat ggacaacttc ttcgcccccg ttttcaccat
     3301 gggcaaatat tatacgcaag gcgacaaggt gctgatgccg ctggcgattc aggttcatca
     3361 tgccgtt
//
