LOCUS       sequence-210090-        4863 bp    DNA     circular SYN 15-APR-2025
DEFINITION  PUPsp1 GFP reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4863)
  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 4863)
  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: "Nat 
            Biotechnol. 2018 Mar 19. pii: nbt.4111. doi: 10.1038/nbt.4111."
COMMENT     Sequence Label: pTHSSe_31
FEATURES             Location/Qualifiers
     source          1..4863
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_RNA        73..122
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     CDS             join(165..167,168..881)
                     /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             165..878
                     /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(195..223)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     818..838
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      898..969
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      985..1012
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_feature    1019..1039
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      1040..1097
                     /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(1121..1138)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(1769..2719)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITLDSYLHNGLRKTLHDALTAKIQITSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      2767..2989
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(3481..3575)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(join(3606..4397,4398..4466))
                     /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     4229..4248
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      complement(4791..4834)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_feature    4837..4858
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
ORIGIN
        1 gaaaatttat caaaaagagt attgacttat aactcaacct ataggatact tacagccatc
       61 gagagctgcg agctgtcacc ggatgtgctt tccggtctga tgagtccgtg aggacgaaac
      121 agcctctaca aataattttg tttaatcaca caggaaagta ctagatgagc aaaggagaag
      181 aacttttcac tggagttgtc ccaattcttg ttgaattaga tggtgatgtt aatgggcaca
      241 aattttctgt ccgtggagag ggtgaaggtg atgctacaaa cggaaaactc acccttaaat
      301 ttatttgcac tactggaaaa ctacctgttc cgtggccaac acttgtcact actctgacct
      361 atggtgttca atgcttttcc cgttatccgg atcacatgaa acggcatgac tttttcaaga
      421 gtgccatgcc cgaaggttat gtacaggaac gcactatatc tttcaaagat gacgggacct
      481 acaagacgcg tgctgaagtc aagtttgaag gtgataccct tgttaatcgt atcgagttaa
      541 agggtattga ttttaaagaa gatggaaaca ttcttggaca caaactcgag tacaacttta
      601 actcacacaa tgtatacatc acggcagaca aacaaaagaa tggaatcaaa gctaacttca
      661 aaattcgcca caacgttgaa gatggttccg ttcaactagc agaccattat caacaaaata
      721 ctccaattgg cgatggccct gtccttttac cagacaacca ttacctgtcg acacaatctg
      781 tcctttcgaa agatcccaac gaaaagcgtg accacatggt ccttcttgag tttgtaactg
      841 ctgctgggat tacacatggc atggatgagc tctacaaata atactagagc caggcatcaa
      901 ataaaacgaa aggctcagtc gaaagactgg gcctttcgtt ttatctgttg tttgtcggtg
      961 aacgctctct actagagtca cactggctca ccttcgggtg ggcctttctg cgtttatata
     1021 ctagtagcgg ccgctgcagt ccggcaaaaa agggcaaggt gtcaccaccc tgcccttttt
     1081 ctttaaaacc gaaaagatta cttcgcgtta tgcaggcttc ctcgctcact gactcgctgc
     1141 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atctgttgtt
     1201 tgtcggtgaa cgctctcctg agtaggacaa atccgccgcc ctagacctag ggtacgggtt
     1261 ttgctgcccg caaacgggct gttctggtgt tgctagtttg ttatcagaat cgcagatccg
     1321 gcttcagccg gtttgccggc tgaaagcact atttcttcca gaattgccat gatttttccc
     1381 cacgggaggc gtcactggct cccgtgttgt cggcagcttt gattcgataa gcagcatcgc
     1441 ctgtttcagg ctgtctatgt gtgactgttg agctgtaaca agttgtctca ggtgttcaat
     1501 ttcatgttct agttgctttg ttttactggt ttcacctgtt ctattaggtg ttacatgctg
     1561 ttcatctgtt acattgtcga tctgttcatg gtgaacagct ttgaatgcac caaaaactcg
     1621 taaaagctct gatgtatcta tcttttttac accgttttca tctgtgcata tggacagttt
     1681 tccctttgat atgtaacggt gaacagttgt tctacttttg tttgttagtc ttgatgcttc
     1741 actgatagat acaagagcca taagaacctc agatccttcc gtatttagcc agtatgttct
     1801 ctagtgtggt tcgttgtttt tgcgtgagcc atgagaacga accattgaga tcatacttac
     1861 tttgcatgtc actcaaaaat tttgcctcaa aactggtgat ctgaattttt gcagttaaag
     1921 catcgtgtag tgtttttctt agtccgttat gtaggtagga atctaatgta atggttgttg
     1981 gtattttgtc accattcatt tttatctggt tgttctcaag ttcggttacg agatccattt
     2041 gtctatctag ttcaacttgg aaaatcaacg tatcagtcgg gcggcctcgc ttatcaacca
     2101 ccaatttcat attgctgtaa gtgtttaaat ctttacttat tggtttcaaa acccattggt
     2161 taagcctttt aaactcatgg tagttatttt caagcattaa catgaactta aattcatcaa
     2221 ggctaatctc tatatttgcc ttgtgagttt tcttttgtgt tagttctttt aataaccact
     2281 cataaatcct catagagtat ttgttttcaa aagacttaac atgttccaga ttatatttta
     2341 tgaatttttt taactggaaa agataaggca atatctcttc actaaaaact aattctaatt
     2401 tttcgcttga gaacttggca tagtttgtcc actggaaaat ctcaaagcct ttaaccaaag
     2461 gattcctgat ttccacagtt ctcgtcatca gctctctggt tgctttagct aatacaccat
     2521 aagcattttc cctactgatg ttcatcatct gagcgtattg gttataagtg aacgataccg
     2581 tccgttcttt ccttgtaggg ttttcaatcg tggggttgag tagtgccaca cagcataaaa
     2641 ttagcttggt ttcatgctcc gttaagtcat agcgactaat cgctagttca tttgctttga
     2701 aaacaactaa ttcagacata catctcaatt ggtctaggtg attttaatca ctataccaat
     2761 tgagatgggc tagtcaatga taattactag tccttttcct ttgagttgtg ggtatctgta
     2821 aattctgcta gacctttgct ggaaaacttg taaattctgc tagaccctct gtaaattccg
     2881 ctagaccttt gtgtgttttt tttgtttata ttcaagtggt tataatttat agaataaaga
     2941 aagaataaaa aaagataaaa agaatagatc ccagccctgt gtataactca ctactttagt
     3001 cagttccgca gtattacaaa aggatgtcgc aaacgctgtt tgctcctcta caaaacagac
     3061 cttaaaaccc taaaggctta agtagcaccc tcgcaagctc gggcaaatcg ctgaatattc
     3121 cttttgtctc cgaccatcag gcacctgagt cgctgtcttt ttcgtgacat tcagttcgct
     3181 gcgctcacgg ctctggcagt gaatgggggt aaatggcact acaggcgcct tttatggatt
     3241 catgcaagga aactacccat aatacaagaa aagcccgtca cgggcttctc agggcgtttt
     3301 atggcgggtc tgctatgtgg tgctatctga ctttttgctg ttcagcagtt cctgccctct
     3361 gattttccag tctgaccact tcggattatc ccgtgacagg tcattcagac tggctaatgc
     3421 acccagtaag gcagcggtat catcaacagg cttacccgtc ttactgtccc tagtgcttgg
     3481 attctcacca ataaaaaacg cccggcggca accgagcgtt ctgaacaaat ccagatggag
     3541 ttctgaggtc attactggat ctatcaacag gagtccaagc gagctctcga accccagagt
     3601 cccgcttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     3661 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     3721 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     3781 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     3841 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     3901 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     3961 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     4021 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     4081 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     4141 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     4201 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     4261 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     4321 agatccagtt cgatataacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     4381 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     4441 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     4501 cgcgaaacga tcctcatcct gtctcttgat cagatcttga tcccctgcgc catcagatcc
     4561 ttggcggcaa gaaagccatc cagtttactt tgcagggctt cccaacctta ccagagggcg
     4621 ccccagctgg caattccgac gtctaagaaa ccattattat catgacatta acctataaaa
     4681 ataggcgtat cacgaggccc tttcgtcttc acctcgagtg ccacctgacg tctaagaaac
     4741 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggcaga atttcagata
     4801 aaaaaaatcc ttagctttcg ctaaggatga tttctggaat tcgcggccgc ttctagagat
     4861 ccc
//
