LOCUS       sequence-95183-e        4367 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PT3 GFP reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4367)
  AUTHORS   Segall-Shapiro TH, Meyer AJ, Ellington AD, Sontag ED, Voigt CA
  TITLE     A 'resource allocator' for transcription based on a highly 
            fragmented T7 RNA polymerase.
  JOURNAL   Mol Syst Biol. 2014 Jul 30;10(7):742.
  PUBMED    25080493
REFERENCE   2  (bases 1 to 4367)
  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: "Mol Syst 
            Biol."; date: "2014-07-30"; volume: "10(7)"; pages: "742"
COMMENT     Sequence Label: pTHSSd_9
FEATURES             Location/Qualifiers
     source          1..4367
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             join(59..61,62..775)
                     /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             59..772
                     /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(89..117)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     712..732
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      796..843
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1418..2368)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLRNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKHDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      2416..2638
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(3130..3224)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(join(3255..4046,4047..4115))
                     /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     3878..3897
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     primer_bind     4329..4347
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 taataaccct cactataggg agaggatcct gtcaatttcc gcgatagagg aggtaaagat
       61 gagcaaagga gaagaacttt tcactggagt tgtcccaatt cttgttgaat tagatggtga
      121 tgttaatggg cacaaatttt ctgtccgtgg agagggtgaa ggtgatgcta caaacggaaa
      181 actcaccctt aaatttattt gcactactgg aaaactacct gttccgtggc caacacttgt
      241 cactactctg acctatggtg ttcaatgctt ttcccgttat ccggatcaca tgaaacggca
      301 tgactttttc aagagtgcca tgcccgaagg ttatgtacag gaacgcacta tatctttcaa
      361 agatgacggg acctacaaga cgcgtgctga agtcaagttt gaaggtgata cccttgttaa
      421 tcgtatcgag ttaaagggta ttgattttaa agaagatgga aacattcttg gacacaaact
      481 cgagtacaac tttaactcac acaatgtata catcacggca gacaaacaaa agaatggaat
      541 caaagctaac ttcaaaattc gccacaacgt tgaagatggt tccgttcaac tagcagacca
      601 ttatcaacaa aatactccaa ttggcgatgg ccctgtcctt ttaccagaca accattacct
      661 gtcgacacaa tctgtccttt cgaaagatcc caacgaaaag cgtgaccaca tggtccttct
      721 tgagtttgta actgctgctg ggattacaca tggcatggat gagctctaca aataataagg
      781 tgcttaataa ggatcctagc ataacccctt ggggcctcta aacgggtctt gaggggtttt
      841 ttgctgttgt ttgtcggtga acgctctcct gagtaggaca aatccgccgc cctagaccta
      901 gggtacgggt tttgctgccc gcaaacgggc tgttctggtg ttgctagttt gttatcagaa
      961 tcgcagatcc ggcttcaggt ttgccggctg aaagcgctat ttcttccaga attgccatga
     1021 ttttttcccc acgggaggcg tcactggctc ccgtgttgtc ggcagctttg attcgataag
     1081 cagcatcgcc tgtttcaggc tgtctatgtg tgactgttga gctgtaacaa gttgtctcag
     1141 gtgttcaatt tcatgttcta gttgctttgt tttactggtt tcacctgttc tattaggtgt
     1201 tacatgctgt tcatctgtta cattgtcgat ctgttcatgg tgaacagctt taaatgcacc
     1261 aaaaactcgt aaaagctctg atgtatctat cttttttaca ccgttttcat ctgtgcatat
     1321 ggacagtttt ccctttgata tctaacggtg aacagttgtt ctacttttgt ttgttagtct
     1381 tgatgcttca ctgatagata caagagccat aagaacctca gatccttccg tatttagcca
     1441 gtatgttctc tagtgtggtt cgttgttttt gcgtgagcca tgagaacgaa ccattgagat
     1501 catgcttact ttgcatgtca ctcaaaaatt ttgcctcaaa actggtgagc tgaatttttg
     1561 cagttaaagc atcgtgtagt gtttttctta gtccgttacg taggtaggaa tctgatgtaa
     1621 tggttgttgg tattttgtca ccattcattt ttatctggtt gttctcaagt tcggttacga
     1681 gatccatttg tctatctagt tcaacttgga aaatcaacgt atcagtcggg cggcctcgct
     1741 tatcaaccac caatttcata ttgctgtaag tgtttaaatc tttacttatt ggtttcaaaa
     1801 cccattggtt aagcctttta aactcatggt agttattttc aagcattaac atgaacttaa
     1861 attcatcaag gctaatctct atatttgcct tgtgagtttt cttttgtgtt agttctttta
     1921 ataaccactc ataaatcctc atagagtatt tgttttcaaa agacttaaca tgttccagat
     1981 tatattttat gaattttttt aactggaaaa gataaggcaa tatctcttca ctaaaaacta
     2041 attctaattt ttcgcttgag aacttggcat agtttgtcca ctggaaaatc tcaaagcctt
     2101 taaccaaagg attcctgatt tccacagttc tcgtcatcag ctctctggtt gctttagcta
     2161 atacaccata agcattttcc ctactgatgt tcatcatctg agcgtattgg ttataagtga
     2221 acgataccgt ccgttctttc cttgtagggt tttcaatcgt ggggttgagt agtgccacac
     2281 agcataaaat tagcttggtt tcatgctccg ttaagtcata gcgactaatc gctagttcat
     2341 ttgctttgaa aacaactaat tcagacatac atctcaattg gtctaggtga ttttaatcac
     2401 tataccaatt gagatgggct agtcaatgat aattactagt ccttttcctt tgagttgtgg
     2461 gtatctgtaa attctgctag acctttgctg gaaaacttgt aaattctgct agaccctctg
     2521 taaattccgc tagacctttg tgtgtttttt ttgtttatat tcaagtggtt ataatttata
     2581 gaataaagaa agaataaaaa aagataaaaa gaatagatcc cagccctgtg tataactcac
     2641 tactttagtc agttccgcag tattacaaaa ggatgtcgca aacgctgttt gctcctctac
     2701 aaaacagacc ttaaaaccct aaaggcttaa gtagcaccct cgcaagctcg ggcaaatcgc
     2761 tgaatattcc ttttgtctcc gaccatcagg cacctgagtc gctgtctttt tcgtgacatt
     2821 cagttcgctg cgctcacggc tctggcagtg aatgggggta aatggcacta caggcgcctt
     2881 ttatggattc atgcaaggaa actacccata atacaagaaa agcccgtcac gggcttctca
     2941 gggcgtttta tggcgggtct gctatgtggt gctatctgac tttttgctgt tcagcagttc
     3001 ctgccctctg attttccagt ctgaccactt cggattatcc cgtgacaggt cattcagact
     3061 ggctaatgca cccagtaagg cagcggtatc atcaacaggc ttacccgtct tactgtccct
     3121 agtgcttgga ttctcaccaa taaaaaacgc ccggcggcaa ccgagcgttc tgaacaaatc
     3181 cagatggagt tctgaggtca ttactggatc tatcaacagg agtccaagcg agctctcgaa
     3241 ccccagagtc ccgcttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     3301 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     3361 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     3421 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     3481 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     3541 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     3601 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     3661 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     3721 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     3781 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     3841 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     3901 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     3961 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     4021 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     4081 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     4141 agcatttatc gcgaaacgat cctcatcctg tctcttgatc agatcttgat cccctgcgcc
     4201 atcagatcct tggcggcaag aaagccatcc agtttacttt gcagggcttc ccaaccttac
     4261 cagagggcgc cccagctggc aattccgacg tctaagaaac cattattatc atgacattaa
     4321 cctataaaaa taggcgtatc acgaggccct ttcgtcttca cctcgag
//
