LOCUS       sequence-206002-        3238 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  p15A BbsI cloning vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3238)
  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 3238)
  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_51
FEATURES             Location/Qualifiers
     source          1..3238
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        21..55
                     /label=BBa_J23105
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.24"
     misc_RNA        63..112
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     CDS             join(155..157,158..871)
                     /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             155..868
                     /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(185..213)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     808..828
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      888..959
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      975..1002
                     /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."
     rep_origin      1212..1757
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     1658..1677
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2116..2907,2908..2976))
                     /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     2739..2758
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2977..3081)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3149..3167
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(3171..3214)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_feature    3217..3238
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
ORIGIN
        1 ggaggtgtct tcggcgcgcc tttacggcta gctcagtcct aggtactatg ctagcaaggt
       61 agctgtcacc ggatgtgctt tccggtctga tgagtccgtg aggacgaaac agcctctaca
      121 aataattttg tttaatcaca caggaaagta ctagatgagc aaaggagaag aacttttcac
      181 tggagttgtc ccaattcttg ttgaattaga tggtgatgtt aatgggcaca aattttctgt
      241 ccgtggagag ggtgaaggtg atgctacaaa cggaaaactc acccttaaat ttatttgcac
      301 tactggaaaa ctacctgttc cgtggccaac acttgtcact actctgacct atggtgttca
      361 atgcttttcc cgttatccgg atcacatgaa acggcatgac tttttcaaga gtgccatgcc
      421 cgaaggttat gtacaggaac gcactatatc tttcaaagat gacgggacct acaagacgcg
      481 tgctgaagtc aagtttgaag gtgataccct tgttaatcgt atcgagttaa agggtattga
      541 ttttaaagaa gatggaaaca ttcttggaca caaactcgag tacaacttta actcacacaa
      601 tgtatacatc acggcagaca aacaaaagaa tggaatcaaa gctaacttca aaattcgcca
      661 caacgttgaa gatggttccg ttcaactagc agaccattat caacaaaata ctccaattgg
      721 cgatggccct gtccttttac cagacaacca ttacctgtcg acacaatctg tcctttcgaa
      781 agatcccaac gaaaagcgtg accacatggt ccttcttgag tttgtaactg ctgctgggat
      841 tacacatggc atggatgagc tctacaaata atactagagc caggcatcaa ataaaacgaa
      901 aggctcagtc gaaagactgg gcctttcgtt ttatctgttg tttgtcggtg aacgctctct
      961 actagagtca cactggctca ccttcgggtg ggcctttctg cgtttataga agacatcgct
     1021 tactagtagc ggccgctgca gtccggcaaa aaagggcaag gtgtcaccac cctgcccttt
     1081 ttctttaaaa ccgaaaagat tacttcgcgt tatgcaggct tcctcgctca ctgactcgct
     1141 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatttaata
     1201 agatgatctt cttgagatcg ttttggtctg cgcgtaatct cttgctctga aaacgaaaaa
     1261 accgccttgc agggcggttt ttcgaaggtt ctctgagcta ccaactcttt gaaccgaggt
     1321 aactggcttg gaggagcgca gtcaccaaaa cttgtccttt cagtttagcc ttaaccggcg
     1381 catgacttca agactaactc ctctaaatca attaccagtg gctgctgcca gtggtgcttt
     1441 tgcatgtctt tccgggttgg actcaagacg atagttaccg gataaggcgc agcggtcgga
     1501 ctgaacgggg ggttcgtgca tacagtccag cttggagcga actgcctacc cggaactgag
     1561 tgtcaggcgt ggaatgagac aaacgcggcc ataacagcgg aatgacaccg gtaaaccgaa
     1621 aggcaggaac aggagagcgc acgagggagc cgccaggggg aaacgcctgg tatctttata
     1681 gtcctgtcgg gtttcgccac cactgatttg agcgtcagat ttcgtgatgc ttgtcagggg
     1741 ggcggagcct atggaaaaac ggctttgccg cggccctctc acttccctgt taagtatctt
     1801 cctggcatct tccaggaaat ctccgccccg ttcgtaagcc atttccgctc gccgcagtcg
     1861 aacgaccgag cgtagcgagt cagtgagcga ggaagcggaa tatatcctgt atcacatatt
     1921 ctgctgacgc accggtgcag ccttttttct cctgccacat gaagcacttc actgacaccc
     1981 tcatcagtgc caacatagta agccagtata cactccgcta cgattatcaa aaaggatctt
     2041 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     2101 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     2161 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     2221 cttaccatct ggccccagtg ctgcaatgat accgcgagaa ccacgctcac cggctccaga
     2281 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     2341 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     2401 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     2461 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     2521 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     2581 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     2641 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     2701 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     2761 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     2821 accgctgttg agatccagtt cgatataacc cactcgtgca cccaactgat cttcagcatc
     2881 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     2941 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     3001 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     3061 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac
     3121 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggcaga atttcagata
     3181 aaaaaaatcc ttagctttcg ctaaggatga tttctggaat tcgcggccgc ttctagag
//
