LOCUS       sequence-210092-        4864 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PUPsp2 GFP reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4864)
  AUTHORS   Segall-Shapiro TH, Sontag ED, Voigt CA
  TITLE     Engineered promoters enable constant gene expression at any copy 
            number in bacteria.
  JOURNAL   Nat Biotechnol. 2018 Mar 19. pii: nbt.4111. doi: 10.1038/nbt.4111.
  PUBMED    29553576
REFERENCE   2  (bases 1 to 4864)
  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 
            Biotechnol. 2018 Mar 19. pii: nbt.4111. doi: 10.1038/nbt.4111."
COMMENT     Sequence Label: pTHSSe_32
FEATURES             Location/Qualifiers
     source          1..4864
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_RNA        75..124
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     CDS             join(167..169,170..883)
                     /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             167..880
                     /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(197..225)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     820..840
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      900..971
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      987..1014
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_feature    1021..1041
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      1042..1099
                     /label=his operon terminator
                     /note="This putative transcriptin terminator from the E. 
                     coli his operon has a 2-bp deletion introduced during 
                     synthesis. Its efficiency has not been determined."
     primer_bind     complement(1123..1140)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(1772..2722)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQITSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      2770..2992
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(3484..3578)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(join(3609..4400,4401..4469))
                     /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     4232..4251
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      complement(4794..4837)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_feature    4840..4861
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
ORIGIN
        1 ccgaaaattt atcaaaaaga gtattgactt atattgagtc gtataggata cttacagcca
       61 tcgagagctg cgagctgtca ccggatgtgc tttccggtct gatgagtccg tgaggacgaa
      121 acagcctcta caaataattt tgtttaatca cacaggaaag tactagatga gcaaaggaga
      181 agaacttttc actggagttg tcccaattct tgttgaatta gatggtgatg ttaatgggca
      241 caaattttct gtccgtggag agggtgaagg tgatgctaca aacggaaaac tcacccttaa
      301 atttatttgc actactggaa aactacctgt tccgtggcca acacttgtca ctactctgac
      361 ctatggtgtt caatgctttt cccgttatcc ggatcacatg aaacggcatg actttttcaa
      421 gagtgccatg cccgaaggtt atgtacagga acgcactata tctttcaaag atgacgggac
      481 ctacaagacg cgtgctgaag tcaagtttga aggtgatacc cttgttaatc gtatcgagtt
      541 aaagggtatt gattttaaag aagatggaaa cattcttgga cacaaactcg agtacaactt
      601 taactcacac aatgtataca tcacggcaga caaacaaaag aatggaatca aagctaactt
      661 caaaattcgc cacaacgttg aagatggttc cgttcaacta gcagaccatt atcaacaaaa
      721 tactccaatt ggcgatggcc ctgtcctttt accagacaac cattacctgt cgacacaatc
      781 tgtcctttcg aaagatccca acgaaaagcg tgaccacatg gtccttcttg agtttgtaac
      841 tgctgctggg attacacatg gcatggatga gctctacaaa taatactaga gccaggcatc
      901 aaataaaacg aaaggctcag tcgaaagact gggcctttcg ttttatctgt tgtttgtcgg
      961 tgaacgctct ctactagagt cacactggct caccttcggg tgggcctttc tgcgtttata
     1021 tactagtagc ggccgctgca gtccggcaaa aaagggcaag gtgtcaccac cctgcccttt
     1081 ttctttaaaa ccgaaaagat tacttcgcgt tatgcaggct tcctcgctca ctgactcgct
     1141 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatctgttg
     1201 tttgtcggtg aacgctctcc tgagtaggac aaatccgccg ccctagacct agggtacggg
     1261 ttttgctgcc cgcaaacggg ctgttctggt gttgctagtt tgttatcaga atcgcagatc
     1321 cggcttcagc cggtttgccg gctgaaagca ctatttcttc cagaattgcc atgatttttt
     1381 ccccacggga ggcgtcactg gctcccgtgt tgtcggcagc tttgattcga taagcagcat
     1441 cgcctgtttc aggctgtcta tgtgtgactg ttgagctgta acaagttgtc tcaggtgttc
     1501 aatttcatgt tctagttgct ttgttttact ggtttcacct gttctattag gtgttacatg
     1561 ctgttcatct gttacattgt cgatctgttc atggtgaaca gctttgaatg caccaaaaac
     1621 tcgtaaaagc tctgatgtat ctatcttttt tacaccgttt tcatctgtgc atatggacag
     1681 ttttcccttt gatatgtaac ggtgaacagt tgttctactt ttgtttgtta gtcttgatgc
     1741 ttcactgata gatacaagag ccataagaac ctcagatcct tccgtattta gccagtatgt
     1801 tctctagtgt ggttcgttgt ttttgcgtga gccatgagaa cgaaccattg agatcatact
     1861 tactttgcat gtcactcaaa aattttgcct caaaactggt gatctgaatt tttgcagtta
     1921 aagcatcgtg tagtgttttt cttagtccgt tatgtaggta ggaatctgat gtaatggttg
     1981 ttggtatttt gtcaccattc atttttatct ggttgttctc aagttcggtt acgagatcca
     2041 tttgtctatc tagttcaact tggaaaatca acgtatcagt cgggcggcct cgcttatcaa
     2101 ccaccaattt catattgctg taagtgttta aatctttact tattggtttc aaaacccatt
     2161 ggttaagcct tttaaactca tggtagttat tttcaagcat taacatgaac ttaaattcat
     2221 caaggctaat ctctatattt gccttgtgag ttttcttttg tgttagttct tttaataacc
     2281 actcataaat cctcatagag tatttgtttt caaaagactt aacatgttcc agattatatt
     2341 ttatgaattt ttttaactgg aaaagataag gcaatatctc ttcactaaaa actaattcta
     2401 atttttcgct tgagaacttg gcatagtttg tccactggaa aatctcaaag cctttaacca
     2461 aaggattcct gatttccaca gttctcgtca tcagctctct ggttgcttta gctaatacac
     2521 cataagcatt ttccctactg atgttcatca tctgagcgta ttggttataa gtgaacgata
     2581 ccgtccgttc tttccttgta gggttttcaa tcgtggggtt gagtagtgcc acacagcata
     2641 aaattagctt ggtttcatgc tccgttaagt catagcgact aatcgctagt tcatttgctt
     2701 tgaaaacaac taattcagac atacatctca attggtctag gtgattttaa tcactatacc
     2761 aattgagatg ggctagtcaa tgataattac tagtcctttt cctttgagtt gtgggtatct
     2821 gtaaattctg ctagaccttt gctggaaaac ttgtaaattc tgctagaccc tctgtaaatt
     2881 ccgctagacc tttgtgtgtt ttttttgttt atattcaagt ggttataatt tatagaataa
     2941 agaaagaata aaaaaagata aaaagaatag atcccagccc tgtgtataac tcactacttt
     3001 agtcagttcc gcagtattac aaaaggatgt cgcaaacgct gtttgctcct ctacaaaaca
     3061 gaccttaaaa ccctaaaggc ttaagtagca ccctcgcaag ctcgggcaaa tcgctgaata
     3121 ttccttttgt ctccgaccat caggcacctg agtcgctgtc tttttcgtga cattcagttc
     3181 gctgcgctca cggctctggc agtgaatggg ggtaaatggc actacaggcg ccttttatgg
     3241 attcatgcaa ggaaactacc cataatacaa gaaaagcccg tcacgggctt ctcagggcgt
     3301 tttatggcgg gtctgctatg tggtgctatc tgactttttg ctgttcagca gttcctgccc
     3361 tctgattttc cagtctgacc acttcggatt atcccgtgac aggtcattca gactggctaa
     3421 tgcacccagt aaggcagcgg tatcatcaac aggcttaccc gtcttactgt ccctagtgct
     3481 tggattctca ccaataaaaa acgcccggcg gcaaccgagc gttctgaaca aatccagatg
     3541 gagttctgag gtcattactg gatctatcaa caggagtcca agcgagctct cgaaccccag
     3601 agtcccgctt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     3661 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     3721 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     3781 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     3841 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     3901 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     3961 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     4021 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     4081 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     4141 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     4201 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     4261 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     4321 ttgagatcca gttcgatata acccactcgt gcacccaact gatcttcagc atcttttact
     4381 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     4441 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     4501 tatcgcgaaa cgatcctcat cctgtctctt gatcagatct tgatcccctg cgccatcaga
     4561 tccttggcgg caagaaagcc atccagttta ctttgcaggg cttcccaacc ttaccagagg
     4621 gcgccccagc tggcaattcc gacgtctaag aaaccattat tatcatgaca ttaacctata
     4681 aaaataggcg tatcacgagg ccctttcgtc ttcacctcga gtgccacctg acgtctaaga
     4741 aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc agaatttcag
     4801 ataaaaaaaa tccttagctt tcgctaagga tgatttctgg aattcgcggc cgcttctaga
     4861 gatc
//
