LOCUS       sequence-384240-        4038 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pFA6a derived selection cassette flanked with transcription 
            terminators (tDEG1/tDEG1), 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 4038)
  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 4038)
  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: kanMX6-ins1
FEATURES             Location/Qualifiers
     source          1..4038
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        41..59
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     41..58
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     gene            222..1578
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        222..565
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             566..1375
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(633..652)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      1381..1578
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(1732..1751)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1733..1751)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1838..1855)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2009..2597)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2089..2108)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2768..3559,3560..3628))
                     /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     3391..3410
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3629..3733)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3801..3819
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3857..3879)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3979..3998
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gctacaatta atacataacc ttatgtatca tacacatacg atttaggtga cactatagaa
       61 cgcggccgcc agctgaagct tcgtacgctg caggtcgacg gatccccggg ttaattaaga
      121 aaagaaatat agtcttcaag gttatattat acaggtttat atattattgg cgcgccagat
      181 ctgtttagct tgcctcgtcc ccgccgggtc acccggccag cgacatggag gcccagaata
      241 ccctccttga cagtcttgac gtgcgcagct caggggcatg atgtgactgt cgcccgtaca
      301 tttagcccat acatccccat gtataatcat ttgcatccat acattttgat ggccgcacgg
      361 cgcgaagcaa aaattacggc tcctcgctgc agacctgcga gcagggaaac gctcccctca
      421 cagacgcgtt gaattgtccc cacgccgcgc ccctgtagag aaatataaaa ggttaggatt
      481 tgccactgag gttcttcttt catatacttc cttttaaaat cttgctagga tacagttctc
      541 acatcacatc cgaacataaa caaccatggg taaggaaaag actcacgttt cgaggccgcg
      601 attaaattcc aacatggatg ctgatttata tgggtataaa tgggctcgcg ataatgtcgg
      661 gcaatcaggt gcgacaatct atcgattgta tgggaagccc gatgcgccag agttgtttct
      721 gaaacatggc aaaggtagcg ttgccaatga tgttacagat gagatggtca gactaaactg
      781 gctgacggaa tttatgcctc ttccgaccat caagcatttt atccgtactc ctgatgatgc
      841 atggttactc accactgcga tccccggcaa aacagcattc caggtattag aagaatatcc
      901 tgattcaggt gaaaatattg ttgatgcgct ggcagtgttc ctgcgccggt tgcattcgat
      961 tcctgtttgt aattgtcctt ttaacagcga tcgcgtattt cgtctcgctc aggcgcaatc
     1021 acgaatgaat aacggtttgg ttgatgcgag tgattttgat gacgagcgta atggctggcc
     1081 tgttgaacaa gtctggaaag aaatgcataa gcttttgcca ttctcaccgg attcagtcgt
     1141 cactcatggt gatttctcac ttgataacct tatttttgac gaggggaaat taataggttg
     1201 tattgatgtt ggacgagtcg gaatcgcaga ccgataccag gatcttgcca tcctatggaa
     1261 ctgcctcggt gagttttctc cttcattaca gaaacggctt tttcaaaaat atggtattga
     1321 taatcctgat atgaataaat tgcagtttca tttgatgctc gatgagtttt tctaatcagt
     1381 actgacaata aaaagattct tgttttcaag aacttgtcat ttgtatagtt tttttatatt
     1441 gtagttgttc tattttaatc aaatgttagc gtgatttata ttttttttcg cctcgacatc
     1501 atctgcccag atgcgaagtt aagtgcgcag aaagtaatat catgcgtcaa tcgtatgtga
     1561 atgctggtcg ctatactgct gtcgattcga tactaacgcc gccatccaaa taatatataa
     1621 acctgtataa tataaccttg aagactatat ttcttttcgt ttaaacgagc tcgaattcat
     1681 cgatgatatc agatccacta gtggcctatg cggccgcgga tctgccggtc tccctatagt
     1741 gagtcgtatt aatttcgata agccaggtta acctgcatta atgaatcggc caacgcgcgg
     1801 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     1861 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     1921 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     1981 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     2041 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     2101 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     2161 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     2221 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     2281 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     2341 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     2401 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     2461 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     2521 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     2581 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     2641 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     2701 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     2761 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     2821 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     2881 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     2941 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     3001 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     3061 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     3121 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     3181 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     3241 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     3301 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     3361 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     3421 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     3481 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     3541 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     3601 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     3661 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     3721 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta
     3781 tcatgacatt aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg
     3841 gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt
     3901 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc
     3961 ggggctggct taactatgcg gcatcagagc agattgtact gagagtgcac catatggaca
     4021 tattgtcgtt agaacgcg
//
