LOCUS       sequence-249621-        4625 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  EMMA Kit # 1000000119 extension.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4625)
  TITLE     Lewin Lab Plasmids
REFERENCE   2  (bases 1 to 4625)
  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"
COMMENT     Sequence Label: pRDAX-EMMI-012 Esp3I//(U)-BsaI-SacB-BsaI-(V)//...
FEATURES             Location/Qualifiers
     source          1..4625
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        86..531
                     /gene="sacR"
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             join(532..618,619..1953)
                     /codon_start=1
                     /gene="Bacillus subtilis sacB"
                     /product="secreted levansucrase that renders bacterial 
                     growth sensitive to sucrose"
                     /label=SacB
                     /note="negative selection marker"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     CDS             2051..2710
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2117..2136)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      2954..2985
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     promoter        2986..3088
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             3089..3904
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNGDRVFRLAQAQSRMNNGLVGA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(3162..3181)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      3928..3955
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     rep_origin      complement(3967..4554)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4047..4066)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(4576..4605)
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
ORIGIN
        1 gttacgctgg agtctgaggc tcgtcctgaa tgcgtctctt ggaagagacc tcgcagcgga
       61 cacttcgcga tcctttttaa cccatcacat atacctgccg ttcactatta tttagtgaaa
      121 tgagatatta tgatattttc tgaattgtga ttaaaaaggc aactttatgc ccatgcaaca
      181 gaaactataa aaaatacaga gaatgaaaag aaacagatag attttttagt tctttaggcc
      241 cgtagtctgc aaatcctttt atgattttct atcaaacaaa agaggaaaat agaccagttg
      301 caatccaaac gagagtctaa tagaatgagg tcgaaaagta aatcgcgcgg gtttgttact
      361 gataaagcag gcaagaccta aaatgtgtaa agggcaaagt gtatactttg gcgtcacccc
      421 ttacatattt taggtctttt tttattgtgc gtaactaact tgccatcttc aaacaggagg
      481 gctggaagaa gcagaccgct aacacagtac ataaaaaagg agacatgaac gatgaacatc
      541 aaaaagtttg caaaacaagc aacagtatta acctttacta ccgcactgct ggcaggaggc
      601 gcaactcaag cgtttgcgaa agaaacgaac caaaagccat ataaggaaac atacggcatt
      661 tcccatatta cacgccatga tatgctgcaa atccctgaac agcaaaaaaa tgaaaaatat
      721 caagttcctg aattcgattc gtccacaatt aaaaatatct cttctgcaaa aggcctggac
      781 gtttgggaca gctggccatt acaaaacgct gacggcactg tcgcaaacta tcacggctac
      841 cacatcgtct ttgcattagc cggagatcct aaaaatgcgg atgacacatc gatttacatg
      901 ttctatcaaa aagtcggcga aacttctatt gacagctgga aaaacgctgg ccgcgtcttt
      961 aaagacagcg acaaattcga tgcaaatgat tctatcctaa aagaccaaac acaagaatgg
     1021 tcaggttcag ccacatttac atctgacgga aaaatccgtt tattctacac tgatttctcc
     1081 ggtaaacatt acggcaaaca aacactgaca actgcacaag ttaacgtatc agcatcagac
     1141 agctctttga acatcaacgg tgtagaggat tataaatcaa tctttgacgg tgacggaaaa
     1201 acgtatcaaa atgtacagca gttcatcgat gaaggcaact acagctcagg cgacaaccat
     1261 acgctgagag atcctcacta cgtagaagat aaaggccaca aatacttagt atttgaagca
     1321 aacactggaa ctgaagatgg ctaccaaggc gaagaatctt tatttaacaa agcatactat
     1381 ggcaaaagca catcattctt ccgtcaagaa agtcaaaaac ttctgcaaag cgataaaaaa
     1441 cgcacggctg agttagcaaa cggcgctctc ggtatgattg agctaaacga tgattacaca
     1501 ctgaaaaaag tgatgaaacc gctgattgca tctaacacag taacagatga aattgaacgc
     1561 gcgaacgtct ttaaaatgaa cggcaaatgg tacctgttca ctgactcccg cggatcaaaa
     1621 atgacgattg acggcattac gtctaacgat atttacatgc ttggttatgt ttctaattct
     1681 ttaactggcc catacaagcc gctgaacaaa actggccttg tgttaaaaat ggatcttgat
     1741 cctaacgatg taacctttac ttactcacac ttcgctgtac ctcaagcgaa aggaaacaat
     1801 gtcgtgatta caagctatat gacaaacaga ggattctacg cagacaaaca atcaacgttt
     1861 gcgccaagct tcctgctgaa catcaaaggc aagaaaacat ctgttgtcaa agacagcatc
     1921 cttgaacaag gacaattaac agttaacaaa taaaaacgca aaagaaaatg ccgatatcct
     1981 attggcattt tcttttattt cttatcaaca taaaggtgaa tcccataggg caggagctaa
     2041 ggaagctaaa atggagaaaa aaatcactgg atataccacc gttgatatat cccaatggca
     2101 tcgtaaagaa cattttgagg catttcagtc agttgctcaa tgtacctata accagaccgt
     2161 tcagctggat attacggcct ttttaaagac cgtaaagaaa aataagcaca agttttatcc
     2221 ggcctttatt cacattcttg cccgcctgat gaatgctcat ccggaattcc gtatggcaat
     2281 gaaagacggt gagctggtga tatgggatag tgttcaccct tgttacaccg ttttccatga
     2341 gcaaactgaa acgttttcat cgctctggag tgaataccac gacgatttcc ggcagtttct
     2401 acacatatat tcgcaagatg tggcgtgtta cggtgaaaac ctggcctatt tccctaaagg
     2461 gtttattgag aatatgtttt tcgtgtcagc caatccctgg gtgagtttca ccagttttga
     2521 tttaaacgtg gccaatatgg acaacttctt cgcccccgtt ttcaccatgg gcaaatatta
     2581 tacgcaaggc gacaaggtgc tgatgccgct ggcgattcag gttcatcatg ccgtttgtga
     2641 tggcttccat gtcggcagaa tgcttaatga attacaacag tactgcgatg agtggcaggg
     2701 cggggcgtaa ttcccggcca ggacccaacg ctgcccgaga tgcgccgcgt gcggctgctg
     2761 gagatggcgg acgcgatgga tatgttctgc caagggttgg tttgcgcatt cacagttctc
     2821 cgcaagaatt gattggctcc aattcttgga gtggtgaatc cgttagcgag gtgccgccgg
     2881 cttccatgct cggaaagccg cacgaggtct ctgttgagag acgtttaaat catacctgac
     2941 ctccatagca gaaagtcaaa agcctccgac cggaggcttt tgacttgatc ggcacgtaag
     3001 aggttccaac tttcaccata atgaaataag atcactaccg ggcgtatttt ttgagttatc
     3061 gagattttca ggagctaagg aagctaaaat gagccatatt caacgggaaa cgtcttgctc
     3121 gaggccgcga ttaaattcca acatggatgc tgatttatat gggtataaat gggctcgcga
     3181 taatgtcggg caatcaggtg cgacaatcta tcgattgtat gggaagcccg atgcgccaga
     3241 gttgtttctg aaacatggca aaggtagcgt tgccaatgat gttacagatg agatggtcag
     3301 gctaaactgg ctgacggaat ttatgcctct tccgaccatc aagcatttta tccgtactcc
     3361 tgatgatgca tggttactca ccactgcgat cccagggaaa acagcattcc aggtattaga
     3421 agaatatcct gattcaggtg aaaatattgt tgatgcgctg gcagtgttcc tgcgccggtt
     3481 gcattcgatt cctgtttgta attgtccttt taacggcgat cgcgtatttc gcctcgctca
     3541 ggcgcaatca cgaatgaata acggtttggt tggtgcgagt gattttgatg acgagcgtaa
     3601 tggctggcct gttgaacaag tctggaaaga aatgcataag cttttgccat tctcaccgga
     3661 ttcagtcgtc actcatggtg atttctcact tgataacctt atttttgacg aggggaaatt
     3721 aataggttgt attgatgttg gacgagtcgg aatcgcagac cgataccagg atcttgccat
     3781 cctatggaac tgcctcggtg agttttctcc ttcattacag aaacggcttt ttcaaaaata
     3841 tggtattgat aatcctgata tgaataaatt gcagtttcac ttgatgctcg atgagttttt
     3901 ctaatgaggg cccaaatgta atcacctggc tcaccttcgg gtgggccttt ctgcgttgct
     3961 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgat gctcaagtca
     4021 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     4081 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     4141 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     4201 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     4261 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     4321 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     4381 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     4441 agttacctcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     4501 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     4561 cctttgattt tctaccgaag aaaggcccac ccgtgaaggt gagccagtga gttgattgca
     4621 gtcca
//
