LOCUS       sequence-147731-        3858 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Supplemental Figure 4. Promoter long ppTEF1. Expresses GFP..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3858)
  AUTHORS   Perez-Pinera P, Han N, Cleto S, Cao J, Purcell O, Shah KA, Lee K, 
            Ram R, Lu TK
  TITLE     Synthetic biology and microbioreactor platforms for programmable 
            production of biologics at the point-of-care.
  JOURNAL   Nat Commun. 2016 Jul 29;7:12211. doi: 10.1038/ncomms12211.
  PUBMED    27470089
REFERENCE   2  (bases 1 to 3858)
  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: "Nat 
            Commun."; date: "2016-07-29"; volume: "7:12211. doi"; pages: " 
            10.1038/ncomms12211"
COMMENT     Sequence Label: PP152
FEATURES             Location/Qualifiers
     source          1..3858
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             54..428
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     terminator      494..741
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(510..528)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     rep_origin      complement(816..1404)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(896..915)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    1524..1573
                     /label=Bxb1 attB
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="attB site for the Mycopbacteriophage Bxb1 integrase 
                     (Xu et al., 2013)"
     CDS             join(2339..2341,2342..2344,2345..3055)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2339..2341,2342..2344,2345..3055)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2384..2405)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2645..2664)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2992..3013
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3080..3109
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             3125..3142
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      3166..3412
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     promoter        3427..3838
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     promoter        join(3846..3858,1..35)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
ORIGIN
        1 catcggcata gtatatcggc atagtataat acgacaaggt gaggaactaa accatggcca
       61 agttgaccag tgccgttccg gtgctcaccg cgcgcgacgt cgccggagcg gtcgagttct
      121 ggaccgaccg gctcgggttc tcccgggact tcgtggagga cgacttcgcc ggtgtggtcc
      181 gggacgacgt gaccctgttc atcagcgcgg tccaggacca ggtggtgccg gacaacaccc
      241 tggcctgggt gtgggtgcgc ggcctggacg agctgtacgc cgagtggtcg gaggtcgtgt
      301 ccacgaactt ccgggacgcc tccgggccgg ccatgaccga gatcggcgag cagccgtggg
      361 ggcgggagtt cgccctgcgc gacccggccg gcaactgcgt gcacttcgtg gccgaggagc
      421 aggactgaca cgtccgacgg cggcccacgg gtcccaggcc tcggagatcc gtcccccttt
      481 tcctttgtcg atatcatgta attagttatg tcacgcttac attcacgccc tccccccaca
      541 tccgctctaa ccgaaaagga aggagttaga caacctgaag tctaggtccc tatttatttt
      601 tttatagtta tgttagtatt aagaacgtta tttatatttc aaatttttct tttttttctg
      661 tacagacgcg tgtacgcatg taacattata ctgaaaacct tgcttgagaa ggttttggga
      721 cgctcgaagg ctttaatttg caagctggag accaacatgt gagcaaaagg ccagcaaaag
      781 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
      841 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
      901 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
      961 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca atgctcacgc
     1021 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     1081 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     1141 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     1201 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca
     1261 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     1321 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     1381 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     1441 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gatcagatct gagctcgttt
     1501 ggccgtggcc gtgctcgtcc tcgtcggccg gcttgtcgac gacggcggtc tccgtcgtca
     1561 ggatcatccg ggccacaagc ttgctgacag aagcctcaag aaaaaaaaaa ttcttcttcg
     1621 actatgctgg aggcagagat gatcgagccg gtagttaact atatatagct aaattggttc
     1681 catcacctcg agtcagcatc tggttacgta actctggcaa ccagtaacac gcttaaggtt
     1741 tggaacaaca ctaaactacc ttgcggtact accattgaca ctacacatcc ttaattccaa
     1801 tcctgtctgg cctccttcac cttttaacca tcttgcccat tccaactcgt gtcagattgc
     1861 gtatcaagtg aaaaaaaaaa aattttaaaa tctttaaccc aatcaggtaa taactgtcgc
     1921 ctcttttatc tgccgcactg catgaggtgt ccccttagtg ggaaagagta ctgagccaac
     1981 cctggaggac agcaagggaa aaatacctac aacttgcttc ataatggtcg taaaaacaat
     2041 ccttgtcgga tataagtgtt gtagactgtc ccttatcctc tgcgatgttc ttcctctcaa
     2101 agtttgcgat ttctctctat cagaattgcc atcaagagac tcaggactaa tttcgcagtc
     2161 ccacacgcac tcgtacatga ttggctgaaa tttccctaaa gaatttcttt ttcacgaaaa
     2221 ttttttttta cacaagattt tcagcagata taaaatggag agcaggacct ccgctgtgac
     2281 tcttcttttt tttcttttat tctcactaca tacattttag ttattcgcca acgaattcat
     2341 ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg
     2401 cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg
     2461 caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct
     2521 cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca
     2581 gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt
     2641 caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt
     2701 gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa
     2761 gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg
     2821 catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga
     2881 ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta
     2941 cctgagcacc cagagcgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct
     3001 gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaaggccgc
     3061 ggcggccgcc agctttctag aacaaaaact catctcagaa gaggatctga atagcgccgt
     3121 cgaccatcat catcatcatc attgaccggt cttgctagat tctaatcaag aggatgtcag
     3181 aatgccattt gcctgagaga tgcaggcttc atttttgata cttttttatt tgtaacctat
     3241 atagtatagg attttttttg tcattttgtt tcttctcgta cgagcttgct cctgatcagc
     3301 ctatctcgca gctgatgaat atcttgtggt aggggtttgg gaaaatcatt cgagtttgat
     3361 gtttttcttg gtatttccca ctcctcttca gagtacagaa gattaagtga gaccttcgtt
     3421 tgtgcggatc ccccacacac catagcttca aaatgtttct actccttttt tactcttcca
     3481 gattttctcg gactccgcgc atcgccgtac cacttcaaaa cacccaagca cagcatacta
     3541 aattttccct ctttcttcct ctagggtgtc gttaattacc cgtactaaag gtttggaaaa
     3601 gaaaaaagag accgcctcgt ttctttttct tcgtcgaaaa aggcaataaa aatttttatc
     3661 acgtttcttt ttcttgaaat tttttttttt agtttttttc tctttcagtg acctccattg
     3721 atatttaagt taataaacgg tcttcaattt ctcaagtttc agtttcattt ttcttgttct
     3781 attacaactt tttttacttc ttgttcatta gaaagaaagc atagcaatct aatctaaggg
     3841 gcggtgttga caattaat
//
