LOCUS       sequence-201705-        4486 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  GoldenBraid vector pDGB2-alpha-2, adapted for S. cerevisiae genome 
            integration by flanking the GB cassette with recombination arms of 
            homology for integration at the YORW-Delta-22 solo LTR loci.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 4486)
  AUTHORS   Perez-Gonzalez A, Kniewel R, Veldhuizen M, Verma HK, 
            Navarro-Rodriguez M, Rubio LM, Caro E
  TITLE     Adaptation of the GoldenBraid modular cloning system and creation of
            a toolkit for the expression of heterologous proteins in yeast 
            mitochondria.
  JOURNAL   BMC Biotechnol. 2017 Nov 13;17(1):80. doi: 
            10.1186/s12896-017-0393-y.
  PUBMED    29132331
REFERENCE   2  (bases 1 to 4486)
  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: "BMC 
            Biotechnol."; date: "2017-11-13"; volume: "17(1):80. doi"; pages: " 
            10.1186/s12896-017-0393-y"
COMMENT     Sequence Label: YORW-Delta-22-alpha-2
FEATURES             Location/Qualifiers
     source          1..4486
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     terminator      complement(22..49)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(73..888)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(73..888)
                     /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
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     796..815
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     CDS             complement(2140..2463)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     primer_bind     2145..2164
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2358..2380
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2372..2389
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2373..2389
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    2390..2446
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(2459..2475)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2459..2475)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2472..2494)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2483..2499
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(2507..2513,2514..2531,2532..2537))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2552..2573
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(3655..4243)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3735..3754)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctatggaaa aacgccagca acgcagaaag gcccacccga aggtgagcca ggtgattatg
       61 cattctaggt gattagaaaa actcatcgag catcaaatga aactgcaatt tattcatatc
      121 aggattatca ataccatatt tttgaaaaag ccgtttctgt aatgaaggag aaaactcacc
      181 gaggcagttc cataggatgg caagatcctg gtatcggtct gcgattccga ctcgtccaac
      241 atcaatacaa cctattaatt tcccctcgtc aaaaataagg ttatcaagtg agaaatcacc
      301 atgagtgacg actgaatccg gtgagaatgg caaaagttta tgcatttctt tccagacttg
      361 ttcaacaggc cagccattac gctcgtcatc aaaatcactc gcatcaacca aaccgttatt
      421 cattcgtgat tgcgcctgag cgagtcgaaa tacgcgatcg ctgttaaaag gacaattaca
      481 aacaggaatc gaatgcaacc ggcgcaggaa cactgccagc gcatcaacaa tattttcacc
      541 tgaatcagga tattcttcta atacctggaa tgctgttttc cctgggatcg cagtggtgag
      601 taaccatgca tcatcaggag tacggataaa atgcttgatg gtcggaagag gcataaattc
      661 cgtcagccag tttagtctga ccatctcatc tgtaacatca ttggcaacgc tacctttgcc
      721 atgtttcaga aacaactctg gcgcatcggg cttcccatac aatcggtaga ttgtcgcacc
      781 tgattgcccg acattatcgc gagcccattt atacccatat aaatcagcat ccatgttgga
      841 atttaatcgc ggccttgagc aagacgtttc ccgttgaata tggctcataa cagaacttat
      901 tatttccttc ctcttttcta cagtatttaa agatacccca agaagctaat tataacaaga
      961 cgaactccaa ttcgcggccg ctagggataa cagggtaata cccactgccc tgaagtccag
     1021 caggtactga ggcttggatc tcaaaagcgt ggggtgcgtg ggcgtgcgct gctcgaggca
     1081 ttttagccgc tgatagtgct ggctccaggg gccactgcgc ttgccatacc cgcacacggg
     1141 gttctttgag gcacttaatt gaccggagat ttaaacgata ttgtcgtcgg ggattagaag
     1201 acattactgg tggggaaccc ttgatgataa taggaaatat atgcgcagta tgctccatcg
     1261 ttacgagatg ttagcagcct cagagcatcc taatgggaat atcaatgcat acacaccgga
     1321 gcttggatat gataaacgaa atattcttga atcgtgagat cgcctgtttt caaaaccgtt
     1381 ggaggcagaa acaattttgt cacaagatgg gcattctacc ccatccttgc tgtattattg
     1441 tagtctcgct ttcttttatg ctggacaaat gagactactg cacattttta tacgttcttg
     1501 gtttttttta aaggtgtggt ttcggcatta tcctgccgca cgtttcttgg ataattcatc
     1561 ctgattctct attttaaacg cttcagccta tcaggatttg gttttgatac atactgcaag
     1621 agtgtatctc gggaacagtc atttattccg caacaaactt aattgcggaa cgcgttaggc
     1681 gatttctagc atatatcaaa taccgttcgc gatttcttct gggttcgtct attttctttt
     1741 aaatacttat taacgtactc aaacaactac acttcgttgt atctcagaat gagatccctc
     1801 agtatgacaa tacatcattc taaacgttcg taaaacacat atgaaacaac tttataacaa
     1861 agcgaacaaa atgggcaaca tgagatgaaa ctccgcgtcc cttagctgaa ctacccaaac
     1921 gtacgaatgc ctgaacaatt agtttagatc cgagattccg cgcttccatc atttagtata
     1981 atccatattt tatataatat ataggataag taacagcccg cgaaaagctt cgtctcagtc
     2041 aggagagaga cccacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg
     2101 cgcgtcagcg ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat cagagcagat
     2161 tgtactgaga gtgcaccata tgcggtgtga aataccgcac agatgcgtaa ggagaaaata
     2221 ccgcatcagg cgccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg
     2281 ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg
     2341 ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg aattcgagct
     2401 cggtacccgg ggatcctcta gagtcgacct gcaggcatgc aagcttggcg taatcatggt
     2461 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     2521 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     2581 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgggg tctctcgctt
     2641 gagacgaagc ttggaccaac tatcatccgc taattactga cattaccaaa tgagatctgt
     2701 gaatgggcaa gataaaaaac aaaaattgaa atgtttgacg ttatgtaaaa ctattaattc
     2761 cttcgctttc ggcggtcaca gaatttgcgt gtagctgact cttgttcaat caatatcatt
     2821 tgttacttta tttgaaagtc tgtattactg cgcctattgt catccgtacc aaagaacgtc
     2881 aaaaagaaac aagataattt ttgtgcttac accatttata gatcactgag cccagaatat
     2941 cgctggagct cagtgtaagt ggcatgaaca caactctgac tgatcgcaca tattgccgtt
     3001 atcataaata ctagttgtac ttgtcaatgc gacgaatggc atcatgccta ttattacgtt
     3061 cctctttttc cgtttcatgt ttccagaatg ctattgaatc taacacttca attataaaaa
     3121 agaataaatc cgcaataatt ttaggctaat tgttgtactg tcaagcgaac ctaatggtta
     3181 aaattcagag gaaccttcga cgtagtctga tcgctacttc tatatcttat gttcccagtc
     3241 aatcaaaagt tgatactata atagctgcca tttatacctg ttagttatgg cgatcgttta
     3301 tcacgacaag cccaattttg gtcgagatgc agccagttgc catctattgt ttccaaagca
     3361 ttcgtttgat tcgtatatga taggaaaaca aattaaacga tgtatcaatt tgctcattca
     3421 tcataaattg tgtttgaggc attactaaaa gagcagcaga tatcagtctg ttggaataaa
     3481 aatcaactat catctactaa ctagtattta cgttactagt atattatcat atacggtgtt
     3541 agaagatgac gcaaatgatg agaaatagtc atctaaatta gtggaagctg aaacgcaagg
     3601 attgataatg taatagggat aacagggtaa tgcggccgcc gcgttgctgg cgtttttcca
     3661 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     3721 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     3781 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     3841 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     3901 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     3961 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     4021 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     4081 cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     4141 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     4201 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     4261 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgca
     4321 ttctaggtga ttagaaaaac tcatcgagca tcaaatgaaa ctgcaattta ttcatatcag
     4381 gattatcaat accatatttt tgaaaaagcc gtttctgtaa tgaaggagaa aactcaccga
     4441 ggcagttcca taggatggca agatcctggt atcggtctgc gattcc
//
