LOCUS       sequence-360837-        7253 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Vector containing a yeast integration cassette for RPL18B-driven 
            expression of PRC1-Link-Neon(gfp), a marker for the vacuole.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7253)
  AUTHORS   Bean BDM, Whiteway M, Martin VJJ
  TITLE     The MyLo CRISPR-Cas9 Toolkit: A Markerless Yeast Localization and 
            Overexpression CRISPR-Cas9 Toolkit.
  JOURNAL   G3 (Bethesda). 2022 Jun 16. pii: 6609175. doi: 
            10.1093/g3journal/jkac154.
  PUBMED    35708612
REFERENCE   2  (bases 1 to 7253)
  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: "G3 
            (Bethesda). 2022 Jun 16. pii: 6609175. doi: 
            10.1093/g3journal/jkac154."
COMMENT     Sequence Label: pBBK60 pHomL-RPL18Bp-PRC1-Link-Neon(gfp)-SSA1t...
FEATURES             Location/Qualifiers
     source          1..7253
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(1..792,793..861))
                     /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     624..643
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(862..966)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      1069..1657
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1558..1577
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     regulatory      5406..5415
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
       61 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
      121 cagtgctgca atgataccgc gggacccacg ctcaccggct ccagatttat cagcaataaa
      181 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
      241 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
      301 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
      361 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
      421 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
      481 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
      541 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
      601 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
      661 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
      721 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
      781 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
      841 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
      901 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
      961 tccgcgcaca tttccccgaa aagtgccacc tgtcatgacc aaaatccctt aacgtgagtt
     1021 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     1081 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     1141 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     1201 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     1261 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     1321 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     1381 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     1441 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     1501 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     1561 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     1621 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     1681 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
     1741 ttctgtggat aaccgtgcgg ccgccaatgt tatctattgg cagatgttca tacggggctt
     1801 tcaaatattg atgaaattat gtgatgttta gaagaagatt cgaactgttt tcagtagatt
     1861 tggtaactgt gcaaccataa ctcatgcccc ctatttatcg gcattgagat tgtaaaggaa
     1921 gtaaaacgca tttttgcaga tctgttctct tatgtatttt taatcgtcct tgtatggaag
     1981 tatcaaaggg gacgttcttc acctccttgg aaaacgcaaa ctttcttttt gcctatcaca
     2041 gtatcttatc gagctaacta ttttcgacac acatgaaaaa gcagaaatat taacgaaaaa
     2101 gaaaagaaag accatgtcat gtacgggcaa tcagaatatg caagatgagc gggaacgggc
     2161 gaaatgtgca cgcgcacaac ttacgaaacg cggatgagtc actgacagcc accgcagagg
     2221 ttctgactcc tactgagctc tattggaggt ggcagaaccg gtaatcccac agtgagcaac
     2281 ttgacaaacc aacagatata ggctgtgtct taaaatatac catccaatac cttgatgaac
     2341 ttttcaaaga tgactaaaat aagtgaaatt tcaacattaa cttcgagctg taaacacagt
     2401 ggtttctttg cataaacacc atcagcctca agtcgtcaag taaagatttc gtgttcatgc
     2461 agatagataa caatctatat gttgataatt agcgttgcct catcaatgcg tacacgaaga
     2521 atatactaaa aaatgagcag gcaagataaa cgaaggcaaa gatgacagag cagaaagccc
     2581 tagtaaagcg tattacaaat gaaaccaaga ttcagattgc gatccctgaa ttcgcatcta
     2641 gatggtagag ccacaaacag ccggtacaag caacgatctc caggaccatc tgaatcatgc
     2701 gcggatgaca cgaactcacg acggcgatca cagacattaa cccacagtac agacactgcg
     2761 acaacgtggc aattcgtcgc aataccgtct cactgaactg gccgataatt gcagacgaac
     2821 gaagaggatg tccaatattt tttttaagga ataaggatac ttcaagacta gattcccccc
     2881 tgcattccca tcagaaccgt aaaccttggc gctttccttg ggaagtattc aagaagtgcc
     2941 ttgtccggtt tctgtggctc acaaaccagc gcgcccgata tggctttctt ttcacttatg
     3001 aatgtaccag tacgggacaa ttagaacgct cctgtaacaa tctctttgca aatgtggggt
     3061 tacattctaa ccatgtcaca ctgctgacga aattcaaagt aaaaaaaaat gggaccacgt
     3121 cttgagaacg atagattttc tttattttac attgaacagt cgttgtctca gcgcgcttta
     3181 tgttttcatt catacttcat attataaaat aacaaaagaa gaatttcata ttcacgccca
     3241 agaaatcagg ctgctttcca aatgcaattg acacttcatt agccatcaca caaaactctt
     3301 tcttgctgga gcttctttta aaaaagacct cagtacacca aacacgttac ccgacctcgt
     3361 tattttacga caactatgat aaaattctga agaaaaaata aaaaaatttt catacttctt
     3421 gcttttattt aaaccattga atgatttctt ttgaacaaaa ctacctgttt caccaaagga
     3481 aatagaaaga aaaaatcaat tagaagaaaa caaaaaacaa aagatctatg aaagcattca
     3541 ccagtttact atgtggacta ggcctgtcca ctacactcgc taaggccatc tcattgcaaa
     3601 gaccgttggg tctagataag gacgttttgc tgcaagctgc ggaaaaattt ggtttggacc
     3661 tcgacctgga tcatctcttg aaggagttgg actccaatgt attggacgct tgggcccaaa
     3721 tagagcattt gtacccaaac caggttatga gccttgaaac ttccactaag ccaaaattcc
     3781 ctgaagcaat caaaacgaag aaagactggg actttgtggt caagaatgac gcaattgaaa
     3841 actatcagct tcgtgtcaac aagattaagg accctaaaat cctgggcatt gacccaaatg
     3901 tcacacagta cacgggttac ttggatgtgg aagacgagga caagcatttc ttcttttgga
     3961 cttttgaaag tagaaacgat cctgcaaagg atccggtcat cctttggttg aacgggggtc
     4021 caggttgttc ttcactaacc gggctgttct ttgaattagg accctcatcc attggacctg
     4081 atttgaaacc catcgggaac ccttactctt ggaacagcaa tgccaccgtg atcttccttg
     4141 accagcctgt caacgttggg ttctcgtatt ccgggtcctc aggtgtttcc aacactgtcg
     4201 ccgctggtaa ggatgtctat aacttcttgg agttgttctt cgatcagttc cctgaatacg
     4261 tcaacaaggg ccaagatttc cacatcgctg gggaatccta cgccggccat tacatccctg
     4321 tttttgcctc tgaaattttg tctcacaagg acagaaactt caacttaacc tccgtcttga
     4381 tcggaaatgg cctcactgac ccattgactc agtataacta ttacgaacca atggcctgtg
     4441 gtgaaggtgg cgaaccatct gttttgccct cggaggaatg ctctgctatg gaagactctt
     4501 tggaacgttg tttgggcttg atcgagtcgt gctatgactc gcaatccgtc tggtcctgtg
     4561 ttccagctac catttattgt aataacgccc aattggctcc ttaccaacgt accggcagaa
     4621 acgtttacga tatcaggaag gattgtgaag gtggcaattt gtgctaccca acgttacaag
     4681 atatcgacga ctacttaaac caggactacg tcaaagaagc tgtcggtgcg gaggttgacc
     4741 actacgaatc ctgtaacttc gatatcaaca gaaatttcct gtttgcgggt gattggatga
     4801 agccttacca caccgccgta acagatcttt tgaatcaaga cctacccatt ctggtatatg
     4861 caggcgataa agatttcatc tgtaactggt tgggtaataa ggcgtggacg gatgtcttgc
     4921 catggaagta cgacgaagaa tttgcaagcc aaaaagtacg taactggact gcttctatca
     4981 ccgacgaggt cgctggtgaa gtcaaatcct acaagcactt cacctatttg agagtcttca
     5041 atggtggcca catggttcca tttgacgtcc ctgaaaacgc cttaagtatg gttaacgaat
     5101 ggatccacgg tggtttctcc ttaggatccg gtccttctgg cccaactggc atggtttcta
     5161 agggtgaaga agataacatg gcttctttgc cagccaccca cgagttgcac atcttcggtt
     5221 ccattaacgg tgtcgacttc gatatggtcg gtcaaggtac cggtaaccca aacgacggtt
     5281 acgaagaatt gaacttgaag tctaccaagg gcgacttgca gttctcccct tggattttgg
     5341 ttccacacat cggttacggt ttccaccaat acttgccata cccagacggt atgtccccat
     5401 tccaagccgc catggttgac ggttctggtt accaagtcca cagaaccatg caattcgaag
     5461 atggtgcttc cttgaccgtg aactacagat acacctacga aggttctcac atcaagggcg
     5521 aagcccaagt taagggtacc ggtttccctg ctgacggtcc agtcatgact aactccttga
     5581 ccgctgctga ctggtgtcgt tccaagaaaa cctacccaaa cgataagacc atcatttcca
     5641 ctttcaagtg gtcctacacc accggtaacg gcaagagata ccgctctacc gctcgtacca
     5701 cctacacctt cgccaagcca atggctgcca actacttgaa gaaccagcct atgtacgttt
     5761 tccgtaagac cgagctgaag cattccaaga ctgaattgaa cttcaaggaa tggcaaaagg
     5821 ctttcaccga cgtcatgggt atggacgaat tgtacaagta actcgagtgg cgccaattgg
     5881 tgcggcaatt gataataacg aaaatgtctt ttaatgatct gggtataatg aggaattttc
     5941 cgaacgtttt tactttatat atatatatac atgtaacata tattctatac gctatagaga
     6001 aaggaaattt ttcaattaaa aaaaaaatag agaaagagtt tcacttcttg attatcgcta
     6061 acactaatgg ttgaagtact gctactttaa ttttatgctg ccaatgagac gacggggtca
     6121 tcacggctca tcatgcgcca aacaaatgtg tgcaatacac gctcggatga ctgcatgatg
     6181 accgcactga ctggggacag cagatccacc taagcctgtg agagaagcag acacccgaca
     6241 gatcaaggca gttaactagt gcactgcagt acaaatcgtc cccaacaaaa gtgggctctc
     6301 aaaattcatc acatttaaat gcatatagga agagcaacag ttggtttgca tctgatgttc
     6361 cttaaagatt tcgacataat gtgcgaagta gatcccttca gcactgtact tgtttttgcg
     6421 actagattgt aaatcattct ttatttaatc tctttcttta actactgctt aaagtataat
     6481 ttggtccgta gtttaataac tatactaagc gaacgctctc tcgaaatttt tcttaacgcg
     6541 tccttgtact gcgtctaacg cttttgccac ttggatttct attataggaa atagtctcac
     6601 ttactgggcg acgaattttc gcgttttgat gaagcacagg aagaatttct ttatgctcag
     6661 aagtttgaca gcaagcaagt tcatcattcg aactagcctt attgttttag ttcagtgaca
     6721 gcgaactgcc gtactcgatg ctttatttct cacggtagag cggaagaaca gatagatccc
     6781 tacatattgc taccacttct attacacaat agtttcaata gcttgcagcg tagctaaact
     6841 ctaaaattta tctaaatcac tcatataaac cgaacccttc cccttccgct tatagtacag
     6901 ctgatgctcc ttcactattt taacatgtgg aattcttgaa agaatgaaat cgccatgcca
     6961 agccatcaca cggtctttta tgcaattgat tgaccgcctg caacacatag gcagtaaaat
     7021 ttttacaagc agtagcagaa caggccacac aatcgcaagt gattaacgtc cacacaggta
     7081 tagggtttct ggaccatatg atacatgctc tggccaagca ttccggctgg tcgctaatcg
     7141 ttgagtgccg agcggccgcg attatcaaaa aggatcttca cctagatcct tttaaattaa
     7201 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cag
//
