LOCUS       sequence-125504-        8316 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses mOrange2 (codon adapted for Streptococcus pneumoniae); 
            under control of Zinc inducible promoter PZn. Plasmid integrates in 
            S. pneumoniae bgaA locus by double cross-over..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8316)
  AUTHORS   Beilharz K, van Raaphorst R, Kjos M, Veening JW
  TITLE     Red fluorescent proteins for gene expression and protein 
            localization studies in Streptococcus pneumoniae and efficient 
            transformation with Gibson assembled DNA.
  JOURNAL   Appl Environ Microbiol. 2015 Aug 7. pii: AEM.02033-15.
  PUBMED    26253684
REFERENCE   2  (bases 1 to 8316)
  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: "Appl 
            Environ Microbiol. 2015 Aug 7. pii: AEM.02033-15."
COMMENT     Sequence Label: pKB01_mOrange2
FEATURES             Location/Qualifiers
     source          1..8316
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(2281..2303)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    2389..2529
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     2444..2463
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(2544..2561)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2715..3303)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2795..2814)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3474..4265,4266..4334))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4097..4116
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4335..4439)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4507..4525
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctagcgctat atgcgttgat gcaatttcta tgcgcacccg ttctcggagc actgtccgac
       61 cgctttggcc gccgcccagt cctgctcgct tcgctacttg gagccactat cgactacgcg
      121 atcatggcga ccacacccgt cctgtggatc tatcgatgca tgcagccggt accatcacaa
      181 gcactttggg acgttctccc ttagtgcttt tttgatttct cataggccgg cctgttagtc
      241 atatggacac ttaaggcaaa ttgttcagaa ctgaataaag ctgacgtttt gcttctatcc
      301 tttctttgag ttttagtgga taatgataat gaacaaggtg ttcataaatc tattataaca
      361 agaattccaa tcccgggtaa agcttccagc ggccgcggag gaaaattaat gtctaaggga
      421 gaagaaaata atatggctat catcaaggaa tttatgcgtt ttaaggtccg tatggaaggt
      481 tcagtcaatg gtcacgaatt tgaaattgaa ggtgaaggag aaggtcgtcc atacgaaggt
      541 tttcaaactg ctaaattgaa ggtaaccaaa ggtggaccat taccttttgc atgggatatt
      601 ttgtcaccac agtttacata cggaagtaaa gcatatgtta agcatccagc tgatattcct
      661 gattacttta aattgagttt tcctgaaggt tttaaatggg aacgtgtcat gaattttgaa
      721 gatggtggag ttgtcaccgt aacacaagat tcttcattgc aggatggaga atttatctac
      781 aaggttaaat tgcgtggaac caattttcca tctgatggtc ctgtcatgca aaagaaaaca
      841 atgggttggg aagcaagttc tgaacgtatg taccctgaag atggagccct taaaggtgaa
      901 atcaagatgc gtttgaaact taaggatggt ggacattaca cttctgaagt caagacaact
      961 tacaaggcca aaaagccagt acagcttcct ggagcttaca tcgttggtat caaattggat
     1021 attacatcac acaatgaaga ttacactatc gttgaacaat atgaacgtgc agaaggtcgt
     1081 cactctactg gtggtatgga tgaactttac aagtaataat aaggatccta actcgagaaa
     1141 gaattcatga gactagtcaa ggtcggcaat tctgcagtac taggacgccg ccaagccagc
     1201 ttaaacccag ctcaatgagc tgggtttttt gtttgttaaa aatgaagaag aaactgtgaa
     1261 gcgtatttat agcaaagcac tcaaaagttt accttatggg tgcttttttc gtgctttttt
     1321 gaaaagacaa aaaaaagaac cttgccaagc aagattcttt tgatagcgct atcgctgagc
     1381 gccggtcgct accattacca gttggtctgg tgtcaaaaat aataataacc gggcaggcca
     1441 tgtctgcccg tatttcgcgt aaggaaatcc attatgtact atttctggtg atgaaatcaa
     1501 cgtaacattt aaagctgtca aagccaaagt catgagatgg cgtatggagc gtaaagctga
     1561 caagagcggt gttgcgatga ttgagatgac cttccttgca ccaagtgaat tgcctcaaga
     1621 aagcactcaa tcaaagattc ttgtagatgg aaaagaactt gctgatttcg ctgaaaatcg
     1681 tcaagactat caaattacct ataaaggtca acggccaaaa gtctcagttg aagaaaacaa
     1741 tcaagtagct tcaactgtgg tagatagtgg agaagatagc cttccagtac ttgttcgcct
     1801 cgtttcagaa agtggaaaac aagtcaagga ataccgtatc cagttgacta aggaaaaacc
     1861 agtttctgct gtacaagaag atcttccaaa actcgaattt gttgaaaaag atttggccta
     1921 caagacagtt gagaaaaaag attcaacact gtatctaggt gaaactcgtg tagaacaaga
     1981 aggaaaagtt ggaaaagaac gtatctttac agtgattaat cctgatggaa gtaaggaaga
     2041 aaaactccgt gaagtggtag aagttccgac agaccgcatc gtcttggttg gaaccaaacc
     2101 agtagctcaa gaagctaaaa aaccacaagt gtcagaaaaa gcagatacaa aaccaattga
     2161 ttcaagtgaa gctgatcaaa ctaataaagc ccagttacca aatacaggta gtgcggcaag
     2221 ccaagcagca gtagcagcag gtttagcctg cctcgcgcgt ttcggtgatg acggtgaaaa
     2281 cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag
     2341 cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg cagccatgac
     2401 ccagtcacgt agcgatagcg gagtgtatac tggcttaact atgcggcatc agagcagatt
     2461 gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac
     2521 cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     2581 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     2641 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     2701 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     2761 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     2821 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     2881 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     2941 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     3001 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     3061 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     3121 ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg
     3181 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     3241 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     3301 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     3361 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     3421 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     3481 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     3541 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     3601 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     3661 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     3721 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     3781 gccattgctg caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     3841 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     3901 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     3961 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     4021 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     4081 ccggcgtcaa cacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     4141 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     4201 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     4261 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     4321 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     4381 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     4441 acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc
     4501 tataaaaata ggcgtatcac gaggcccttt cgtcttcaag aattaattcc ttcttaacgc
     4561 cccaagttca tcaccaatga catcaactca catgaactac atgatgaacc cagttatcat
     4621 ggttttggat aagatttttg aaaaattctt cccaggcctt gataaatatg actttgatgc
     4681 tgctaaattg aacaagaaaa tcggtttctg gggatctaaa ttcttcatcg gtttcatcct
     4741 tggtatcgtt atcggtatta tgggaactcc acatccaatt gcaggtgttg cagatgcaga
     4801 taaatggcgt cttgttatca aaggatggtt gtctcttggt ttgactgccg gtgtatcttt
     4861 ggaactcttc tcacttatcg gttcatggtt catcgcagcc gtagaaccac tatcacaagg
     4921 tattacaaac gttgctacta aacgtcttca aggacgtaaa ttcaatatcg gtcttgactg
     4981 gccattcatc gctggtcgtg ctgaaatctg ggcttgtgcc aacgtacttg caccaatcat
     5041 gttgattgaa gcagtgcttc tttcaaaagt tggaaatggt atcttgccac ttgcaggtat
     5101 catcgctatg ggtgttactc cagctctctt ggttgtaact cgtggtaaat tgctccgtat
     5161 gattatcttc ggaacactct tgttgccact cttccttctt tcaggtacac ttattgcacc
     5221 atttgcaaca gaacttgcta aaggtgtagg tgccttccca gaaggtgtga gccaaactca
     5281 attgattact cactctactc ttgaaggacc aatcgaaaaa cttcttggtt ggacaattgg
     5341 taacactaca actggtgata tcaaagcaat ccttggtgca gtagccttcc ttgtattcta
     5401 tatcggtatc tttgcttggt acagaaaaca aatgatcaaa cgtaacgaag agtacgcagc
     5461 aaaagcaaaa taatgcgctc cgctagcttt acagacaaag aactatcctt aatggaagca
     5521 ctagcaaaag gtatcaatga agcaagaaat attgaaggct agtcagtaaa attcagataa
     5581 acaaaaagag ccgataagat gagagtttca tttctcaaat tatcggctct gcgtcttggc
     5641 gtctggctct ttgattgtta ttatattcat tttttgaaca ttaattaaag tttcgttctt
     5701 acatttgggg cagtataaag ggaaattttt aagtatagta tccactcgta ttcgtagtct
     5761 tgttttattt ccacaaatag gacacaatat ccacttgtag tttataataa caatctcctc
     5821 ctttccactt taattcaaat ctatattaaa gaatatttca tcttatttaa taagaaacca
     5881 tatttatata acaacataaa acgcactaag ttattttatt gaacatatat cttactttat
     5941 ctatctgact atttagacga cgggtctggc aaacaggttc gccagtggta acctgatatc
     6001 cttttagctc tgctaaacaa acactaagcc catttgtaaa aaaagttaaa tcattgcgat
     6061 aatcttgaat acatcgagca ggaatttctc caataataat gacctcatta tttttcagtt
     6121 gagtatttac gatatttgca caatatttgg gagcatcgtt atatgcccgt gaaagatatt
     6181 cctgtggtgc ataaacttta aaactaagat atggctctaa caattctgtt ccagcttttc
     6241 taaaggcttg ctccaataca ataggagcaa gcatccgaaa atctgctggg gtactaacag
     6301 ggctatagta taagccatac ttaaaacaga ttttacagtc cgtcacattc caaccataca
     6361 atccttgttc acaaccatag cgtatccctt ccataactgc attttgaaat gattgattta
     6421 agtatccaag agaaaccgag ctctcatact gcattccact tcccaacgga agcggtgata
     6481 cagataaacc aatggaagcc cagaaaggat ttggcggcac ttcgatgtga atggtatatt
     6541 ctgcattttt taacggtctc tccatataaa tgactgtagg ctcttttagt tctatctcca
     6601 catgatactt ttcttgcaac agtgcactaa tcacttccat ttgtactttc cctaagaaag
     6661 aaagtataat ttcatgtgtc gtagaatcca cgtaatatcg tagaagcgga tcactatctg
     6721 agatttccaa aagggcatca agcaacattt ctctctgttc aggtttactc ggttcaacag
     6781 ttgtttgtag tagagggtgc ggattttcaa tcttttttct ctgtggcaat agttttgtat
     6841 ctccaagaac actatttaac ttcaaaaact cattttgcaa aataacaatt tctccagaat
     6901 aagctctatc aatcttacat aattcaccat ttattgaagt atacatttct gtaactttta
     6961 ttttttcttt ttctgatact ctaaccgaat ctcgtaaatg tagtactcca ctataaaggc
     7021 gtatatatgc aagacgttgt cttttttttg tatattcaat tttgaaaaca tttccgcaaa
     7081 gttcagacgg acctcgatgt gttgatgaat aaaatttatt agtaataact tctataaggt
     7141 tatcaatccc tatattactt tttgcacttc catgataaag agggaacaga gaacaattct
     7201 gaaatcttat gctttcctct tgttcgagtt ccaatgcttc taatgattta ccggacatat
     7261 atttctctaa aaggtcatcg tttccctcta ttaccgtatc ccattgttca gattcggtaa
     7321 agttcgtcac acacacatta ggatacagtt ctaccttctg tttgattaca atttcggcag
     7381 aaagtttctc tttaatatcc tgataaaccg ttgataaatc aattccattt tggtcaatct
     7441 tattgataaa aaagattgtg ggaatcccca ttttcctaag tgcatgaaat aatatacgag
     7501 tttgtgcttg tacgccatct tttgcagaaa tcagtagaat tgccccatct aaaactgata
     7561 atgaacgata tacttctgct aagaaatcca tatgtcctgg cgtgtctatg atgttcacct
     7621 tcgtattttc ccactgaaaa gaggttattc ctgtctgaat tgtaattcct ctctgacgtt
     7681 ctaaaagcgt attatccgtc ctcgttgtac ctttgtccac gcttcctaat tctgtaatcg
     7741 ctccactgtt atataataag ctttctgtta aggtagtttt tcctgcatca acatgagcta
     7801 aaactccaat attaataatt ttcatgtgat tttcctccat tcaaaagccc aaaagggcat
     7861 aaaaatccca gtgataaata ctcttatcac tgggattttt atgcataacc ataggcatac
     7921 aaagcataca gatattctcc ggatacttta gaatcacatg ataaaggtat tcttaaactg
     7981 ggtacaaaaa actaagccct cctaaaaaag gacatccaat tatttgttcc cgctatcaaa
     8041 ttgacagttt atttaagaat accttgccgc atatttatta actcctttta aatagatact
     8101 taaattatag cacgtaagag catatttgta aaggaatctc caatttttta tcaaaaagag
     8161 tacatgatta caaagtatct gtaatcatgt accaatattt gttattttac aatcttccaa
     8221 ttatcttgtt tttctaatat caactcaaat tgcgagattt gggttgcctt tgtttcttga
     8281 tctaaatatt tcacagcgac attcacgatt acttgg
//
