LOCUS       sequence-384245-        3973 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 3973)
  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 3973)
  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: natMX6-ins5
FEATURES             Location/Qualifiers
     source          1..3973
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..19
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1..18
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     gene            182..1301
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     promoter        182..525
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             526..1098
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     terminator      1104..1301
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(1373..1391)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(1627..1646)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1628..1646)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1733..1750)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1904..2492)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1984..2003)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2663..3454,3455..3523))
                     /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     3286..3305
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3524..3628)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3696..3714
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3752..3774)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3874..3893
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 atttaggtga cactatagaa cgcggccgcc agctgaagct tcgtacgctg caggtcgacg
       61 gatccccggg ttaattaaga aaagaaatat agtcttcaag gttatattat acaggtttat
      121 atattattgg cgcgccagat ctgtttagct tgcctcgtcc ccgccgggtc acccggccag
      181 cgacatggag gcccagaata ccctccttga cagtcttgac gtgcgcagct caggggcatg
      241 atgtgactgt cgcccgtaca tttagcccat acatccccat gtataatcat ttgcatccat
      301 acattttgat ggccgcacgg cgcgaagcaa aaattacggc tcctcgctgc agacctgcga
      361 gcagggaaac gctcccctca cagacgcgtt gaattgtccc cacgccgcgc ccctgtagag
      421 aaatataaaa ggttaggatt tgccactgag gttcttcttt catatacttc cttttaaaat
      481 cttgctagga tacagttctc acatcacatc cgaacataaa caaccatggg taccactctt
      541 gacgacacgg cttaccggta ccgcaccagt gtcccggggg acgccgaggc catcgaggca
      601 ctggatgggt ccttcaccac cgacaccgtc ttccgcgtca ccgccaccgg ggacggcttc
      661 accctgcggg aggtgccggt ggacccgccc ctgaccaagg tgttccccga cgacgaatcg
      721 gacgacgaat cggacgacgg ggaggacggc gacccggact cccggacgtt cgtcgcgtac
      781 ggggacgacg gcgacctggc gggcttcgtg gtcgtctcgt actccggctg gaaccgccgg
      841 ctgaccgtcg aggacatcga ggtcgccccg gagcaccggg ggcacggggt cgggcgcgcg
      901 ttgatggggc tcgcgacgga gttcgcccgc gagcggggcg ccgggcacct ctggctggag
      961 gtcaccaacg tcaacgcacc ggcgatccac gcgtaccggc ggatggggtt caccctctgc
     1021 ggcctggaca ccgccctgta cgacggcacc gcctcggacg gcgagcaggc gctctacatg
     1081 agcatgccct gcccctaatc agtactgaca ataaaaagat tcttgttttc aagaacttgt
     1141 catttgtata gtttttttat attgtagttg ttctatttta atcaaatgtt agcgtgattt
     1201 atattttttt tcgcctcgac atcatctgcc cagatgcgaa gttaagtgcg cagaaagtaa
     1261 tatcatgcgt caatcgtatg tgaatgctgg tcgctatact gctgtcgatt cgatactaac
     1321 gccgccatcc aacaggcccc ttttcctttg tcgatatcat gtaattagtt atgtcacgct
     1381 tacattcacg ccctcccccc acatccgctc taaccgaaaa ggaaggagtt agacaacctg
     1441 aagtctaggt ccctatttat ttttttatag ttatgttagt attaagaacg ttatttatat
     1501 ttcaaatttt tctttttttt ctgtacaaac gcgtgtacgc atgtaacatt atagtttaaa
     1561 cgagctcgaa ttcatcgatg atatcagatc cactagtggc ctatgcggcc gcggatctgc
     1621 cggtctccct atagtgagtc gtattaattt cgataagcca ggttaacctg cattaatgaa
     1681 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     1741 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     1801 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     1861 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     1921 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     1981 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     2041 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     2101 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     2161 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     2221 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     2281 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     2341 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     2401 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     2461 agcagattac gctcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     2521 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     2581 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     2641 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     2701 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     2761 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     2821 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     2881 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     2941 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     3001 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     3061 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     3121 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     3181 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     3241 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     3301 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     3361 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     3421 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     3481 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     3541 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     3601 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct
     3661 aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc
     3721 gtctcgcgcg tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg
     3781 tcacagcttg tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg
     3841 gtgttggcgg gtgtcggggc tggcttaact atgcggcatc agagcagatt gtactgagag
     3901 tgcaccatat ggacatattg tcgttagaac gcggctacaa ttaatacata accttatgta
     3961 tcatacacat acg
//
