LOCUS       sequence-210100-        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        23..57
                     /label=BBa_J23105
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.24"
     misc_RNA        65..114
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     CDS             join(157..159,160..873)
                     /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             157..870
                     /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(187..215)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     810..830
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      890..961
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      977..1004
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_feature    1023..1043
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      1044..1101
                     /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      1214..1759
                     /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     1660..1679
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2118..2909,2910..2978))
                     /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     2741..2760
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2979..3083)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3151..3169
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(3173..3216)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_feature    join(3219..3238,1..2)
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
ORIGIN
        1 agggaggtgt cttcggcgcg cctttacggc tagctcagtc ctaggtacta tgctagcaag
       61 gtagctgtca ccggatgtgc tttccggtct gatgagtccg tgaggacgaa acagcctcta
      121 caaataattt tgtttaatca cacaggaaag tactagatga gcaaaggaga agaacttttc
      181 actggagttg tcccaattct tgttgaatta gatggtgatg ttaatgggca caaattttct
      241 gtccgtggag agggtgaagg tgatgctaca aacggaaaac tcacccttaa atttatttgc
      301 actactggaa aactacctgt tccgtggcca acacttgtca ctactctgac ctatggtgtt
      361 caatgctttt cccgttatcc ggatcacatg aaacggcatg actttttcaa gagtgccatg
      421 cccgaaggtt atgtacagga acgcactata tctttcaaag atgacgggac ctacaagacg
      481 cgtgctgaag tcaagtttga aggtgatacc cttgttaatc gtatcgagtt aaagggtatt
      541 gattttaaag aagatggaaa cattcttgga cacaaactcg agtacaactt taactcacac
      601 aatgtataca tcacggcaga caaacaaaag aatggaatca aagctaactt caaaattcgc
      661 cacaacgttg aagatggttc cgttcaacta gcagaccatt atcaacaaaa tactccaatt
      721 ggcgatggcc ctgtcctttt accagacaac cattacctgt cgacacaatc tgtcctttcg
      781 aaagatccca acgaaaagcg tgaccacatg gtccttcttg agtttgtaac tgctgctggg
      841 attacacatg gcatggatga gctctacaaa taatactaga gccaggcatc aaataaaacg
      901 aaaggctcag tcgaaagact gggcctttcg ttttatctgt tgtttgtcgg tgaacgctct
      961 ctactagagt cacactggct caccttcggg tgggcctttc tgcgtttata gaagacatcg
     1021 cttactagta gcggccgctg cagtccggca aaaaagggca aggtgtcacc accctgccct
     1081 ttttctttaa aaccgaaaag attacttcgc gttatgcagg cttcctcgct cactgactcg
     1141 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatttaa
     1201 taagatgatc ttcttgagat cgttttggtc tgcgcgtaat ctcttgctct gaaaacgaaa
     1261 aaaccgcctt gcagggcggt ttttcgaagg ttctctgagc taccaactct ttgaaccgag
     1321 gtaactggct tggaggagcg cagtcaccaa aacttgtcct ttcagtttag ccttaaccgg
     1381 cgcatgactt caagactaac tcctctaaat caattaccag tggctgctgc cagtggtgct
     1441 tttgcatgtc tttccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     1501 gactgaacgg ggggttcgtg catacagtcc agcttggagc gaactgccta cccggaactg
     1561 agtgtcaggc gtggaatgag acaaacgcgg ccataacagc ggaatgacac cggtaaaccg
     1621 aaaggcagga acaggagagc gcacgaggga gccgccaggg ggaaacgcct ggtatcttta
     1681 tagtcctgtc gggtttcgcc accactgatt tgagcgtcag atttcgtgat gcttgtcagg
     1741 ggggcggagc ctatggaaaa acggctttgc cgcggccctc tcacttccct gttaagtatc
     1801 ttcctggcat cttccaggaa atctccgccc cgttcgtaag ccatttccgc tcgccgcagt
     1861 cgaacgaccg agcgtagcga gtcagtgagc gaggaagcgg aatatatcct gtatcacata
     1921 ttctgctgac gcaccggtgc agcctttttt ctcctgccac atgaagcact tcactgacac
     1981 cctcatcagt gccaacatag taagccagta tacactccgc tacgattatc aaaaaggatc
     2041 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     2101 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     2161 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     2221 ggcttaccat ctggccccag tgctgcaatg ataccgcgag aaccacgctc accggctcca
     2281 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     2341 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     2401 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     2461 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     2521 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     2581 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     2641 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     2701 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     2761 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     2821 ttaccgctgt tgagatccag ttcgatataa cccactcgtg cacccaactg atcttcagca
     2881 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     2941 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     3001 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     3061 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa
     3121 accattatta tcatgacatt aacctataaa aataggcgta tcacgaggca gaatttcaga
     3181 taaaaaaaat ccttagcttt cgctaaggat gatttctgga attcgcggcc gcttctag
//
