LOCUS       sequence-382613-        7978 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Red marker for yeast late Golgi/early endosome. Integration of 
            2xKatushka tag at SEC7 C terminus. Uses antibiotic resistance marker
            hphMX6. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7978)
  AUTHORS   Li D, Yang SG, He CW, Zhang ZT, Liang Y, Li H, Zhu J, Su X, Gong Q, 
            Xie Z
  TITLE     Excess diacylglycerol at the endoplasmic reticulum disrupts 
            endomembrane homeostasis and autophagy.
  JOURNAL   BMC Biol. 2020 Aug 28;18(1):107. doi: 10.1186/s12915-020-00837-w.
  PUBMED    32859196
REFERENCE   2  (bases 1 to 7978)
  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: "BMC Biol.";
            date: "2020-08-28"; volume: "18(1):107. doi"; pages: " 
            10.1186/s12915-020-00837-w"
COMMENT     Sequence Label: SEC7-2Katushka-ter-Hygromycin
FEATURES             Location/Qualifiers
     source          1..7978
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      2578..2765
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2633..2657)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     protein_bind    2817..2850
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     gene            2905..4480
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        2905..3248
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             3249..4277
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             3249..4277
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     terminator      4283..4480
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    4543..4576
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(5615..5634)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5616..5634)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5721..5738)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5892..6480)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5972..5991)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6651..7442,7443..7511))
                     /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     7274..7293
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7512..7616)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7684..7702
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7740..7762)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7862..7881
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        join(7962..7978,1..2)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     join(7962..7978,1)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 gaacgcggcc gccagctgaa gcttgatatt gcaaaacgta tctaaattca atgagtacca
       61 ttggaatcaa ataggtgacg tattcgataa attgtttgat ttaaccactg ctaatgaatt
      121 gttcgattat gatccgttac agcaaggaag aaaatcatct gtatctcatc accaaacaac
      181 taatgatacc agccaacact cagatgatga tagcaacgac agaagggaga acgattctaa
      241 tatcagtgag acggtcgaaa gagcgcatca agaagaatcc agcgaagacg ttggtggtga
      301 catggttgaa acactcaatg ggcaaactaa gcttaataat ggcaattccg ttccgacggt
      361 aaaggacgaa ttgaatccaa agcccgctag tctaagtatc cccaagaaaa ctaagcacat
      421 gaaacgaaac gaaagtaacg aagatatacg taggagaata aacatcaaga attctattgt
      481 tgtcaaatgt gttttacagc ttttgatgat agagctgctc aacgaattat tcgagaacga
      541 agattttgcc cactgtatcc cttacaagga agccattaga attacaagat tgttggagaa
      601 atcgtacgaa ttttctcgtg actttaatga agattatggg ttaagaacaa gactagtaga
      661 ggctcgtgta gttgataaaa taccaaacct gttgaaacaa gaaacgagtg ctgctgcagt
      721 tctccttgac attatgtttc agttgtatct gaacgatgat gagaagaagg ctgatttaat
      781 aactcgtctg atcaccattt gcatccaggt cgtagagggt tatgtttctt tggatgatag
      841 aaccatggaa cgtagtatca atgcctggcg ttcagtgata gttgaaatat tacaaggtta
      901 ctacgaattt gatgatgaag actttagact atactgtcct gctatgtatg ctctggtgat
      961 tcaaatatta gataagagcg ttccaacgga gttgagacat gctataaaac aatttctaag
     1021 cagagttggt gaattatacc tttctactga tgcttcgtac gctgcaggtc gtgggattcc
     1081 tgggttaatt aacatcgcta gcatggtcgg agaggatagt gtgttaataa ccgaaaatat
     1141 gcacatgaag ctatacatgg agggaaccgt gaatgatcac catttcaaat gcactagcga
     1201 gggcgaagga aagccttatg aaggtacaca aactatgaaa ataaaggtcg ttgaaggggg
     1261 gccactacca ttcgctttcg acatattggc gacctccttt atgtatggaa gtaagacttt
     1321 tatcaaccat actcagggga ttcctgattt ttttaaacaa tcttttccag aaggcttcac
     1381 ctgggagcgt ataacgacct atgaagatgg aggcgtatta acggcgacac aggacacgtc
     1441 actacaaaat ggctgcttga tatataatgt taaaatcaac ggagtcaatt tcccctcaaa
     1501 tggcccggtg atgcagaaga aaacattggg gtgggaagcg tccactgaaa tgttataccc
     1561 ggctgactct ggtttgcgtg gccacgcaca gatggcgcta aagctagtag gaggtggtta
     1621 tctgcactgc tcactaaaaa ctacctatag atcaaagaaa cctgcgaaga accttaaaat
     1681 gccaggattc tattttgtag acagacgtct agagcgtatt aaggaggctg ataaggagac
     1741 ttacgttgaa caacacgaaa tggctgtagc gagatactgc gatttgccct ctaagttagg
     1801 gcatagcatt ccagttccaa cgcgtgatct agagaagcca ttcatggtcg gagaggatag
     1861 tgtgttaata accgaaaata tgcacatgaa gctatacatg gagggaaccg tgaatgatca
     1921 ccatttcaaa tgcactagcg agggcgaagg aaagccttat gaaggtacac aaactatgaa
     1981 aataaaggtc gttgaagggg ggccactacc attcgctttc gacatattgg cgacctcctt
     2041 tatgtatgga agtaagactt ttatcaacca tactcagggg attcctgatt tttttaaaca
     2101 atcttttcca gaaggcttca cctgggagcg tataacgacc tatgaagatg gaggcgtatt
     2161 aacggcgaca caggacacgt cactacaaaa tggctgcttg atatataatg ttaaaatcaa
     2221 cggagtcaat ttcccctcaa atggcccggt gatgcagaag aaaacattgg ggtgggaagc
     2281 gtccactgaa atgttatacc cggctgactc tggtttgcgt ggccacgcac agatggcgct
     2341 aaagctagta ggaggtggtt atctgcactg ctcactaaaa actacctata gatcaaagaa
     2401 acctgcgaag aaccttaaaa tgccaggatt ctattttgta gacagacgtc tagagcgtat
     2461 taaggaggct gataaggaga cttacgttga acaacacgaa atggctgtag cgagatactg
     2521 cgatttgccc tctaagttag ggcatagccc taggtgaggc gcgccacttc taaataagcg
     2581 aatttcttat gatttatgat ttttattatt aaataagtta taaaaaaaat aagtgtatac
     2641 aaattttaaa gtgactctta ggttttaaaa cgaaaattct tattcttgag taactctttc
     2701 ctgtaggtca ggttgctttc tcaggtatag tatgaggtcg ctcttattga ccacacctct
     2761 accggcagat ccgctaggga taacagggta atatctgcag gtcgacaacc cttaatataa
     2821 cttcgtataa tgtatgctat acgaagttat taggtctaga gatctgttta gcttgcctcg
     2881 tccccgccgg gtcacccggc cagcgacatg gaggcccaga ataccctcct tgacagtctt
     2941 gacgtgcgca gctcaggggc atgatgtgac tgtcgcccgt acatttagcc catacatccc
     3001 catgtataat catttgcatc catacatttt gatggccgca cggcgcgaag caaaaattac
     3061 ggctcctcgc tgcagacctg cgagcaggga aacgctcccc tcacagacgc gttgaattgt
     3121 ccccacgccg cgcccctgta gagaaatata aaaggttagg atttgccact gaggttcttc
     3181 tttcatatac ttccttttaa aatcttgcta ggatacagtt ctcacatcac atccgaacat
     3241 aaacaaccat gggtaaaaag cctgaactca ccgcgacgtc tgtcgagaag tttctgatcg
     3301 aaaagttcga cagcgtctcc gacctgatgc agctctcgga gggcgaagaa tctcgtgctt
     3361 tcagcttcga tgtaggaggg cgtggatatg tcctgcgggt aaatagctgc gccgatggtt
     3421 tctacaaaga tcgttatgtt tatcggcact ttgcatcggc cgcgctcccg attccggaag
     3481 tgcttgacat tggggaattc agcgagagcc tgacctattg catctcccgc cgtgcacagg
     3541 gtgtcacgtt gcaagacctg cctgaaaccg aactgcccgc tgttctgcag ccggtcgcgg
     3601 aggcaatgga tgcgatcgct gcggccgatc ttagccagac gagcgggttc ggcccattcg
     3661 gaccgcaagg aatcggtcaa tacactacat ggcgtgattt catatgcgcg attgctgatc
     3721 cccatgtgta tcactggcaa actgtgatgg acgacaccgt cagtgcgtcc gtcgcgcagg
     3781 ctctcgatga gctgatgctt tgggccgagg actgccccga agtccggcac ctcgtgcacg
     3841 cggatttcgg ctccaacaat gtcctgacgg acaatggccg cataacagcg gtcattgact
     3901 ggagcgaggc gatgttcggg gattcccaat acgaggtcgc caacatcttc ttctggaggc
     3961 cgtggttggc ttgtatggag cagcagacgc gctacttcga gcggaggcat ccggagcttg
     4021 caggatcgcc acggctccgg gcgtatatgc tccgcattgg tcttgaccaa ctctatcaga
     4081 gcttggttga cggcaatttc gatgatgcag cttgggcgca gggtcgatgc gacgcaatcg
     4141 tccgatccgg agccgggact gtcgggcgta cacaaatcgc ccgcagaagc gcggccgtct
     4201 ggaccgatgg ctgtgtagaa gtactcgccg atagtggaaa ccgacgcccc agcactcgtc
     4261 cgagggcaaa ggaataatca gtactgacaa taaaaagatt cttgttttca agaacttgtc
     4321 atttgtatag tttttttata ttgtagttgt tctattttaa tcaaatgtta gcgtgattta
     4381 tatttttttt cgcctcgaca tcatctgccc agatgcgaag ttaagtgcgc agaaagtaat
     4441 atcatgcgtc aatcgtatgt gaatgctggt cgctatactg ctgtcgattc gatactaacg
     4501 ccgccatcca gtgtcgaaaa cgagctctcg agaaccctta atataacttc gtataatgta
     4561 tgctatacga agttattagg tgatatcaga tccactagtg gcctatgcgg ccgcggcaga
     4621 ttaaaatatg cttagttgta gaacacagaa aaaatgagaa ggttgatata cttttactca
     4681 tattaatcct gatttagtgg cggtgatcct ttttagattc gtttaattca taaacaaggt
     4741 acatcatata gcaataacaa atatgtaaac tatgtagaaa gacttatcaa gtttagccat
     4801 tttggtcctc attcatgggc attttatact aacttcattt ttgccgaaat aagaaaggat
     4861 gtaatcttaa cctaaaatat gtaagcaatt aagggcgcac gtagtcactt gcgaacaaat
     4921 aggaaaagaa agaaacaagc acggatactg aatgactgag tgtattacta cacgatttat
     4981 ctcaacgaat ttaagtgtgg aggacgatcg gctgcacgta ggagtcaacg tttttttact
     5041 gtgttcatca caaaaccatt aagctgagtc atccatattg caaacaataa ggtagcaccg
     5101 cttccatcaa aacactagaa cataccgatg ttttcacgga tttattggaa acaaatagca
     5161 gatgagggcg tttttacttg taatgaaggc gaatgagaag gcaaggctta caggcttgtt
     5221 gtttatgcca agcatcaata ctcgctatgg aggagcgcca tcagcgataa caacaaagtt
     5281 tgcaagtgct tgcttggata taaaaggctt gacgcttaag ataaataggg ccatacttgc
     5341 ttgcttttac tatttttgtt cctcataagc ctgattgact acgctattat aaacccctac
     5401 attatcctat aatcaacttc acttgtgtaa cgttgaatga gttcaactga gtaatactcg
     5461 gtcggaaata tacgctccat ctggcaggaa acataggctg catttatatc ttgcacaatt
     5521 atgcccttcg taggtgtatg ttttcatgtt ttccattatg tcaatgtaga tgtatctttg
     5581 gacaggcaac ttttggccgc ggatctgccg gtctccctat agtgagtcgt attaatttcg
     5641 ataagccagg ttaacctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     5701 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     5761 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     5821 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     5881 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     5941 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     6001 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     6061 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     6121 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     6181 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     6241 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     6301 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     6361 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     6421 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     6481 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     6541 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     6601 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     6661 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     6721 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     6781 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     6841 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     6901 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     6961 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     7021 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     7081 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     7141 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     7201 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     7261 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     7321 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     7381 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     7441 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     7501 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     7561 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     7621 tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac attaacctat
     7681 aaaaataggc gtatcacgag gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac
     7741 ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc
     7801 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg gcttaactat
     7861 gcggcatcag agcagattgt actgagagtg caccatatgg acatattgtc gttagaacgc
     7921 ggctacaatt aatacataac cttatgtatc atacacatac gatttaggtg acactata
//
