LOCUS       sequence-268963-        6853 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Integration of VPS4-mTagBFP2 as an additional copy in genome, use 
            auxotrophic marker TRP1(Kluyveromyces lactis). Marker for yeast late
            endosome..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6853)
  AUTHORS   Zhu J, Zhang ZT, Tang SW, Zhao BS, Li H, Song JZ, Li D, Xie Z
  TITLE     A Validated Set of Fluorescent-Protein-Based Markers for Major 
            Organelles in Yeast (Saccharomyces cerevisiae).
  JOURNAL   MBio. 2019 Sep 3;10(5). pii: mBio.01691-19. doi: 
            10.1128/mBio.01691-19.
  PUBMED    31481383
REFERENCE   2  (bases 1 to 6853)
  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: "MBio."; 
            date: "2019-09-3"; volume: "10(5). pii: mBio.01691-19. doi"; pages: 
            " 10.1128/mBio.01691-19"
COMMENT     Sequence Label: ZJOM113
FEATURES             Location/Qualifiers
     source          1..6853
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     139..158
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     392..409
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     496..515
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        496..514
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    complement(568..601)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(1604..1637)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     terminator      complement(1689..1876)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     1797..1821
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     promoter        complement(5003..5021)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(5004..5021)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(5102..5121)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5221..5243
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5281..5299)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5367..5471
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5472..5540,5541..6332)
                     /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     complement(5690..5709)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(6503..6853,1..238)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
       61 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
      121 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
      181 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
      241 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
      301 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
      361 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      421 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcaggttaac
      481 ctggcttatc gaaattaata cgactcacta tagggagacc ggcagatccg cggccgcata
      541 ggccactagt ggatctgata tcacctaata acttcgtata gcatacatta tacgaagtta
      601 tattaagggt tctcgagagc tcggtaccgg gcccatgttt gatacatgat tgcatggtat
      661 cactacacac atcccatata ccccagtatc tctcactgaa caaataataa tggtatttta
      721 tgtataaact caaatccttt attctttatt tttgtataat agtaatatca attttaatat
      781 aaggttattt cattgaaaaa ttgaactatc aaacatcagt ttactaatca tttacttggg
      841 tactctgagc gagctaccct cttgcgaaat ctactgtgat gctcgttaaa gtgtgtggtt
      901 tgcaaaccgt tgaagctgca aagactgctg tggatgatgg tgctgattac ttaggtatca
      961 tttgtgttcc cggtaggaaa agaaccattg actcatctgt tgcgaaaggt atttcaactg
     1021 cagttcacca acaagagaac gtgaaaggta ctaaactagt cggggtgttt agaaatcagt
     1081 ccgttgatga tgtccttcaa ctgtaccacg aatataatct agatgtgata caattacatg
     1141 gagatgaaga tattaaagaa tacagatctt tgatcccatc ttcaattcca atcattaaga
     1201 ggttccagtt cccacaggat tgtgaattac tactggacct gtatgaacac gtagacaatg
     1261 tgctgacgtt gttcgattct ggtgaaggtg gcactggtga gaaattgaat tggagtgcaa
     1321 tttccagttg gtctgcaagt catcccgaga taaaattcat tatcgctggt ggattgaatc
     1381 ctgataacgt ttctgttgcc attaatatgt taccaaatgc gatcggtgtc gatgtaagtg
     1441 gaggagtaga gactgatggt atcaaggatt tagaaaaggt aaagctattc atccagcagg
     1501 cctctcaata gaggaaaaaa cgtatctttc ttcaattaca catacgtata tataaattga
     1561 tatatatata gtgaatgatc gttccgcggc cgctctagac ctaataactt cgtatagcat
     1621 acattatacg aagttatatt aagggttgtc gacctgcaga tattaccctg ttatccctag
     1681 cggatctgcc ggtagaggtg tggtcaataa gagcgacctc atactatacc tgagaaagca
     1741 acctgaccta caggaaagag ttactcaaga ataagaattt tcgttttaaa acctaagagt
     1801 cactttaaaa tttgtataca cttatttttt ttataactta tttaataata aaaatcataa
     1861 atcataagaa attcgcttat ttagaagtgg cgcgcctcaa ttcagtttat gacccagctt
     1921 gctaggcaag tcgcaatatc tcgcgactgc aacctcgtgc tgctccacgt aagtctcatt
     1981 atttgcttct ttaattcttt ccagacggta gtcgacatag taaacacccg gcatcttcag
     2041 attttttgcg ggtttcttgc ttctgtaagt ggtcttggca tttgcgatca aatggctgcc
     2101 accaaccagc ttcagtgcca tgtcgttacg gccctccagg ccaccatcgg caggatacag
     2161 ggtttctgta aaggcctccc aacccagggt tttcttctgc atcacgggac cattgctagt
     2221 aaagttgacg ccacgaattt tcacgttata gatcagacag ccatcttgca gggaggtatc
     2281 ttgggtggcg gtcaacacac caccatcttc atacgtggtg acacgctccc aggtgaaacc
     2341 ttccggaaag ctctgtttga agaaatcagg aatgccctga gtatggttaa tgaaggtttt
     2401 ggaaccgtac aggaagctgg tggccagaat gtcaaaagcg aatggcagtg gaccgccctc
     2461 cacaactttg atacgcatgg tttgggtgcc ttcgtagggc ttaccctcac cctcagaggt
     2521 gcacttaaag tggtggttgt caacggtgcc ttccatgtac agcttcatgt gcatgttctc
     2581 tttgatcagt tcgctcattt cttcagcaac ttcaagagaa aggttacctt cttgaccaaa
     2641 atctctagtg aactgttctt gcttcagcaa gtcatcctca ttcacggtag gtctcgtcga
     2701 tttgatagcc tttaagaaat cctttatggt gagatctggt tcttttaatt catcagcttc
     2761 aatatccgtc caactcatct cgatggcgcc atcatcacct ggagagcatg gtgttagttt
     2821 tcttgtttcg tcgtcctccg tagagacatc tttgaaatgt gtcgcgcttt gaatttttct
     2881 tatcggttgc attagcgcat ctttcaccac tacagcaata tcactcccgg aataaccttc
     2941 agtcattgca cctaaagttc tataatcttc cttggtcaat acgcatggag tgtcgccgac
     3001 gttaatctcg aacatggtgg ttcttgccgc caaatctggt aatggtatat atattcttct
     3061 ttcaaatctt ctcctgatgg cactgtccag ttgccatggg atatttgttg cgccaaggac
     3121 caaaacacct tgggagtcat tcccaacacc attcatttgt accaataatt ctgttttaat
     3181 tcttctactt gcttcgctct ctccttctcc tctggtacct gttagcgcat ccacttcatc
     3241 tataaaaata atagaaggtt tattttctct tgccatcgca aataactgtt tgacaagttt
     3301 ttcagattca cccatccact tagaaaccaa atcactggaa ctaacactga aaaaggtaga
     3361 attggcctcg gtggccactg cttttgccaa atatgattta cctgtacctg gtggtccgta
     3421 caataagatt cccgaagtgg gcttacggtt acctttgaac aaatgtggga acttgacagg
     3481 taaaataacg gcttctttaa gggcctcttt ggctccttcc agtccagcca cgtcttccca
     3541 cttaacattg ggcttctcag acaaaatagc actcgataag gcacctctta gctttttgtt
     3601 gtcttctcca ccattatcct caccctcttc ctgagagatt ttcttattac caccgttgga
     3661 accactacca gcgctaggag atttctttgc cgcgtttgcc tcttcacttt ccaagtgctt
     3721 tttcaactgc tctgcacggt ttagatactc tgtaaacttc gctcttatca aatccttcga
     3781 cttagggttt ttctcgtatt tcaaagccaa catcaaataa tctaatccgt tatagtatgc
     3841 tgtatacgcc tcctcatact gcgtggctgt gtccaggtca atggcctttt gaaccagttc
     3901 tattccctta gttaaaaaat ctcccgtgct catttgtacc ccatctacta taggcactct
     3961 atgctgcttt atttttgtct tccatgtcct caacccacga ttaaatcgtt cgatatcatg
     4021 aagttcttat tttttttttt ttttcaccca gcgggggtac ccgtaaagca atacaataaa
     4081 tatcatcaag caagaataaa tagatgtaca gaaagaacaa ccatgacgtg ttgtgtttct
     4141 attcagcgtg cggaaccgaa tcctcagtcg gagattctct gagaatgttc gtttgaggaa
     4201 gcttaggata cttggcgtag aagttggggc cctttggggg ctgtagggaa gaaactggtc
     4261 gttggcgcga ctggagtggc gacatgtgaa tcggacttga agtcactaac gggtttactg
     4321 ctcttactgg cgaaggtggt aaattttctg tcggtttgag gttgtctcgc tcaatatttt
     4381 tcatgtctcg acgagactcc aggtatcttt gatcgatgtt tgcgtcgtcc aacaccgact
     4441 gcaacagatg acacttttcc ttgacaaatt gtagttcctt ttgtgtagag tgcagctgct
     4501 gccgcaattg cgcaaacttt gcttgccatt gtagaggcac acaatcctga gaatttttag
     4561 acaatgggac aacgctttct tcttcctcat ctgcgggaga cccccataca aacgcttgga
     4621 tcttggcaag agtcttagca atattgaatc caaacgcgtg ttgttgagcc ctctttttgg
     4681 agatattctt gtagttttta gaacgtgtcc tttgtgaagc gtgtctaaca tgatttcttt
     4741 cgcgaggcac aaccaccaat ttgcgctctt ttcgtatccc caatgtcttt tcctgaattc
     4801 ccattgtgat cttcaatgcc aacccaacct tctttttact ttgttgggtc taaattatgt
     4861 tactttgtcc atttcatctc ttttcaagat ggaatctaaa agaagaaaaa cgacaaaagg
     4921 aacaaaagtg atggctagcg atgttaatta acccaggaat cccacgacct gcagcgtacg
     4981 aagcttcagc tggcggccgc gttctatagt gtcacctaaa tcgtatgtgt atgatacata
     5041 aggttatgta ttaattgtag ccgcgttcta acgacaatat gtccatatgg tgcactctca
     5101 gtacaatctg ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg
     5161 acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct
     5221 ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg
     5281 gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt
     5341 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     5401 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     5461 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     5521 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     5581 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     5641 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     5701 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     5761 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     5821 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     5881 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     5941 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     6001 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     6061 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     6121 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     6181 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     6241 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     6301 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     6361 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     6421 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     6481 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     6541 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     6601 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     6661 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     6721 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     6781 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     6841 agcccagctt gga
//
