LOCUS       sequence-361034-        5004 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Parent vector for for creating new markerless yeast integration 
            cassettes with different promoters/genes/tags/terminators.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5004)
  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 5004)
  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: pBBK96 pHomL-ConLS-GFPdropout-ConR1-HomR (AmpR)
FEATURES             Location/Qualifiers
     source          1..5004
                     /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"
     RBS             2977..2988
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             2995..3708
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      3720..3791
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3807..3834
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
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 gtgagaccga aagtgaaacg tgatttcatg cgtcattttg aacattttgt aaatcttatt
     2881 taataatgtg tgcggcaatt cacatttaat ttatgaatgt tttcttaaca tcgcggcaac
     2941 tcaagaaacg gcaggttcgg atcttagcta ctagagaaag aggagaaata ctagatgcgt
     3001 aaaggcgaag agctgttcac tggtgtcgtc cctattctgg tggaactgga tggtgatgtc
     3061 aacggtcata agttttccgt gcgtggcgag ggtgaaggtg acgcaactaa tggtaaactg
     3121 acgctgaagt tcatctgtac tactggtaaa ctgccggttc cttggccgac tctggtaacg
     3181 acgctgactt atggtgttca gtgctttgct cgttatccgg accatatgaa gcagcatgac
     3241 ttcttcaagt ccgccatgcc ggaaggctat gtgcaggaac gcacgatttc ctttaaggat
     3301 gacggcacgt acaaaacgcg tgcggaagtg aaatttgaag gcgataccct ggtaaaccgc
     3361 attgagctga aaggcattga ctttaaagag gacggcaata tcctgggcca taagctggaa
     3421 tacaatttta acagccacaa tgtttacatc accgccgata aacaaaaaaa tggcattaaa
     3481 gcgaatttta aaattcgcca caacgtggag gatggcagcg tgcagctggc tgatcactac
     3541 cagcaaaaca ctccaatcgg tgatggtcct gttctgctgc cagacaatca ctatctgagc
     3601 acgcaaagcg ttctgtctaa agatccgaac gagaaacgcg atcatatggt tctgctggag
     3661 ttcgtaaccg cagcgggcat cacgcatggt atggatgaac tgtacaaatg accaggcatc
     3721 aaataaaacg aaaggctcag tcgaaagact gggcctttcg ttttatctgt tgtttgtcgg
     3781 tgaacgctct ctactagagt cacactggct caccttcggg tgggcctttc tgcgtttata
     3841 ggtctcagct gccaatgaga cgacggggtc atcacggctc atcatgcgcc aaacaaatgt
     3901 gtgcaataca cgctcggatg actgcatgat gaccgcactg actggggaca gcagatccac
     3961 ctaagcctgt gagagaagca gacacccgac agatcaaggc agttaactag tgcactgcag
     4021 tacaaatcgt ccccaacaaa agtgggctct caaaattcat cacatttaaa tgcatatagg
     4081 aagagcaaca gttggtttgc atctgatgtt ccttaaagat ttcgacataa tgtgcgaagt
     4141 agatcccttc agcactgtac ttgtttttgc gactagattg taaatcattc tttatttaat
     4201 ctctttcttt aactactgct taaagtataa tttggtccgt agtttaataa ctatactaag
     4261 cgaacgctct ctcgaaattt ttcttaacgc gtccttgtac tgcgtctaac gcttttgcca
     4321 cttggatttc tattatagga aatagtctca cttactgggc gacgaatttt cgcgttttga
     4381 tgaagcacag gaagaatttc tttatgctca gaagtttgac agcaagcaag ttcatcattc
     4441 gaactagcct tattgtttta gttcagtgac agcgaactgc cgtactcgat gctttatttc
     4501 tcacggtaga gcggaagaac agatagatcc ctacatattg ctaccacttc tattacacaa
     4561 tagtttcaat agcttgcagc gtagctaaac tctaaaattt atctaaatca ctcatataaa
     4621 ccgaaccctt ccccttccgc ttatagtaca gctgatgctc cttcactatt ttaacatgtg
     4681 gaattcttga aagaatgaaa tcgccatgcc aagccatcac acggtctttt atgcaattga
     4741 ttgaccgcct gcaacacata ggcagtaaaa tttttacaag cagtagcaga acaggccaca
     4801 caatcgcaag tgattaacgt ccacacaggt atagggtttc tggaccatat gatacatgct
     4861 ctggccaagc attccggctg gtcgctaatc gttgagtgcc gagcggccgc gattatcaaa
     4921 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     4981 atatgagtaa acttggtctg acag
//
