LOCUS       sequence-331130-        2639 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  His-tagged sfGFP for protein purification.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2639)
  TITLE     Sensitizing cell-free molecular detection using cascaded genetic 
            circuitry
REFERENCE   2  (bases 1 to 2639)
  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"
COMMENT     Sequence Label: pJBL7068
FEATURES             Location/Qualifiers
     source          1..2639
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      17..605
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     506..525
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     633..652
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        633..651
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             684..704
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             717..734
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             744..1457
                     /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="SKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGT
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKAN
                     FKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     CDS             744..1454
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Nager et al., 2011)"
                     /label=superfolder GFP
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGT
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKAN
                     FKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     complement(771..799)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1394..1414
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     complement(1640..1658)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1654..1701
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1719..2534)
                     /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="MSHIQRETSCSKPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2442..2461
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2526..2545
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
       61 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
      121 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
      181 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
      241 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
      301 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
      361 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
      421 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
      481 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
      541 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
      601 ggaaaaacgc cagcaacgcg atcccgcgaa attaatacga ctcactatag ggagaccaca
      661 acggtttccc tctagaaata attttgttta actttaagaa ggaggaagga tctatgcatc
      721 accatcacca tcacggctca ggaagcaaag gagaagaact tttcactgga gttgtcccaa
      781 ttcttgttga attagatggt gatgttaatg ggcacaaatt ttctgtccgt ggagagggtg
      841 aaggtgatgc tacaaacgga aaactcaccc ttaaatttat ttgcactact ggaaaactac
      901 ctgttccgtg gccaacactt gtcactactc tgacctatgg tgttcaatgc ttttcccgtt
      961 atccggatca catgaaacgg catgactttt tcaagagtgc catgcccgaa ggttatgtac
     1021 aggaacgcac tatatctttc aaagatgacg ggacctacaa gacgcgtgct gaagtcaagt
     1081 ttgaaggtga tacccttgtt aatcgtatcg agttaaaggg tattgatttt aaagaagatg
     1141 gaaacattct tggacacaaa ctcgagtaca actttaactc acacaatgta tacatcacgg
     1201 cagacaaaca aaagaatgga atcaaagcta acttcaaaat tcgccacaac gttgaagatg
     1261 gttccgttca actagcagac cattatcaac aaaatactcc aattggcgat ggccctgtcc
     1321 ttttaccaga caaccattac ctgtcgacac aatctgtcct ttcgaaagat cccaacgaaa
     1381 agcgtgacca catggtcctt cttgagtttg taactgctgc tgggattaca catggcatgg
     1441 atgagctcta caaataagga tctgaagggg atcccgactg gcgagagcca ggtaacgaat
     1501 ggatcgggtc ggcatggcat ctccacctcc tcgcggtccg acctgggcat ccgaaggagg
     1561 acgtcgtcca ctcggatggc taagggaggt cgaccggctg ctaacaaagc ccgaaaggaa
     1621 gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa
     1681 cgggtcttga ggggtttttt gctgaaagcc aattctgatt agaaaaactc atcgagcatc
     1741 aaatgaaact gcaatttatt catatcagga ttatcaatac catatttttg aaaaagccgt
     1801 ttctgtaatg aaggagaaaa ctcaccgagg cagttccata ggatggcaag atcctggtat
     1861 cggtctgcga ttccgactcg tccaacatca atacaaccta ttaatttccc ctcgtcaaaa
     1921 ataaggttat caagtgagaa atcaccatga gtgacgactg aatccggtga gaatggcaaa
     1981 agcttatgca tttctttcca gacttgttca acaggccagc cattacgctc gtcatcaaaa
     2041 tcactcgcat caaccaaacc gttattcatt cgtgattgcg cctgagcgag acgaaatacg
     2101 cgatcgctgt taaaaggaca attacaaaca ggaatcgaat gcaaccggcg caggaacact
     2161 gccagcgcat caacaatatt ttcacctgaa tcaggatatt cttctaatac ctggaatgct
     2221 gttttcccgg ggatcgcagt ggtgagtaac catgcatcat caggagtacg gataaaatgc
     2281 ttgatggtcg gaagaggcat aaattccgtc agccagttta gtctgaccat ctcatctgta
     2341 acatcattgg caacgctacc tttgccatgt ttcagaaaca actctggcgc atcgggcttc
     2401 ccatacaatc gatagattgt cgcacctgat tgcccgacat tatcgcgagc ccatttatac
     2461 ccatataaat cagcatccat gttggaattt aatcgcggct tcgagcaaga cgtttcccgt
     2521 tgaatatggc tcataacacc ccttgtatta ctgtttatgt aagcagacag ttttattgtt
     2581 catgatgata tatttttatc ttgtgcaatg taacatcaga gattttgaga cacaacgtg
//
