LOCUS       sequence-384243-        4099 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pFA6a derived selection cassette flanked with transcription 
            terminators (tDEG1/tCYC1), allows genome modification without 
            disruption of insertion neighboring genes by transcription 
            interference.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4099)
  AUTHORS   Powers EN, Chan C, Doron-Mandel E, Llacsahuanga Allcca L, Kim Kim J,
            Jovanovic M, Brar GA
  TITLE     Bidirectional promoter activity from expression cassettes can drive 
            off-target repression of neighboring gene translation.
  JOURNAL   Elife. 2022 Dec 12;11:e81086. doi: 10.7554/eLife.81086.
  PUBMED    36503721
REFERENCE   2  (bases 1 to 4099)
  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: "Elife."; 
            date: "2022-12-12"; volume: "11:e81086. doi"; pages: " 
            10.7554/eLife.81086"
COMMENT     Sequence Label: his3MX6-ins3
FEATURES             Location/Qualifiers
     source          1..4099
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        170..513
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             514..1212
                     /codon_start=1
                     /product="S. kluyveri imidazoleglycerol phosphate 
                     dehydratase"
                     /label=k-HIS3
                     /translation="MAEPAQKKQKQTVQERKAFISRITNETKIQIAISLNGGYIQIKDS
                     ILPAKKDDDVASQATQSQVIDIHTGVGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHT
                     TEDCGIALGQAFKEAMGAVRGVKRFGTGFAPLDEALSRAVVDLSSRPFAVIDLGLKREM
                     IGDLSTEMIPHFLESFAEAARITLHVDCLRGFNDHHRSESAFKALAVAIREAISSNGTN
                     DVPSTKGVLM"
     terminator      1218..1415
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(1487..1505)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(1741..1760)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1742..1760)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1847..1864)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2018..2606)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2098..2117)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2777..3568,3569..3637))
                     /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     3400..3419
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3638..3742)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3810..3828
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3866..3888)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3988..4007
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        join(4088..4099,1..7)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     join(4088..4099,1..6)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 ctatagaacg cggccgccag ctgaagcttc gtacgctgca ggtcgacgga tccccgggtt
       61 aattaagaaa agaaatatag tcttcaaggt tatattatac aggtttatat attattggcg
      121 cgccagatct gtttagcttg cctcgtcccc gccgggtcac ccggccagcg acatggaggc
      181 ccagaatacc ctccttgaca gtcttgacgt gcgcagctca ggggcatgat gtgactgtcg
      241 cccgtacatt tagcccatac atccccatgt ataatcattt gcatccatac attttgatgg
      301 ccgcacggcg cgaagcaaaa attacggctc ctcgctgcag acctgcgagc agggaaacgc
      361 tcccctcaca gacgcgttga attgtcccca cgccgcgccc ctgtagagaa atataaaagg
      421 ttaggatttg ccactgaggt tcttctttca tatacttcct tttaaaatct tgctaggata
      481 cagttctcac atcacatccg aacataaaca accatggcag aaccagccca aaaaaagcaa
      541 aaacaaactg ttcaggagcg caaggcgttt atctcccgta tcactaatga aactaaaatt
      601 caaatcgcta tttcgctgaa tggtggttat attcaaataa aagattcgat tcttcctgca
      661 aagaaggatg acgatgtagc ttcccaagct actcagtcac aggtcatcga tattcacaca
      721 ggtgttggct ttttggatca tatgatccat gcgttggcaa aacactctgg ttggtctctt
      781 attgttgaat gtattggtga cctgcacatt gacgatcacc atactaccga agattgcggt
      841 atcgcattag ggcaagcgtt caaagaagca atgggtgctg tccgtggtgt aaaaagattc
      901 ggtactgggt tcgcaccatt ggatgaggcg ctatcacgtg ccgtagtcga tttatctagt
      961 agaccatttg ctgtaatcga ccttggattg aagagagaga tgattggtga tttatccact
     1021 gaaatgattc cacacttttt ggaaagtttc gcggaggcgg ccagaattac tttgcatgtt
     1081 gattgtctga gaggtttcaa cgatcaccac agaagtgaga gtgcgttcaa ggctttggct
     1141 gttgccataa gagaagctat ttctagcaat ggcaccaatg acgttccctc aaccaaaggt
     1201 gttttgatgt gatcagtact gacaataaaa agattcttgt tttcaagaac ttgtcatttg
     1261 tatagttttt ttatattgta gttgttctat tttaatcaaa tgttagcgtg atttatattt
     1321 tttttcgcct cgacatcatc tgcccagatg cgaagttaag tgcgcagaaa gtaatatcat
     1381 gcgtcaatcg tatgtgaatg ctggtcgcta tactgctgtc gattcgatac taacgccgcc
     1441 atccaacagg ccccttttcc tttgtcgata tcatgtaatt agttatgtca cgcttacatt
     1501 cacgccctcc ccccacatcc gctctaaccg aaaaggaagg agttagacaa cctgaagtct
     1561 aggtccctat ttattttttt atagttatgt tagtattaag aacgttattt atatttcaaa
     1621 tttttctttt ttttctgtac aaacgcgtgt acgcatgtaa cattatagtt taaacgagct
     1681 cgaattcatc gatgatatca gatccactag tggcctatgc ggccgcggat ctgccggtct
     1741 ccctatagtg agtcgtatta atttcgataa gccaggttaa cctgcattaa tgaatcggcc
     1801 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     1861 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     1921 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     1981 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     2041 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     2101 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     2161 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     2221 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     2281 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     2341 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     2401 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     2461 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     2521 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     2581 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     2641 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     2701 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     2761 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     2821 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     2881 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     2941 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     3001 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     3061 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     3121 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     3181 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     3241 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     3301 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     3361 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     3421 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     3481 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     3541 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     3601 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     3661 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     3721 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa
     3781 ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc tttcgtctcg
     3841 cgcgtttcgg tgatgacggt gaaaacctct gacacatgca gctcccggag acggtcacag
     3901 cttgtctgta agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg
     3961 gcgggtgtcg gggctggctt aactatgcgg catcagagca gattgtactg agagtgcacc
     4021 atatggacat attgtcgtta gaacgcggct acaattaata cataacctta tgtatcatac
     4081 acatacgatt taggtgaca
//
