LOCUS       sequence-382498-        8021 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Marker for DAG. Yeast expression of mouse PKCdelta under LSY2 
            promoter. Uses antibiotic resistance marker natMX6. Replaces 
            endogenous TRP1 upon genome integration, leading to trp1D. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8021)
  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 8021)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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:  trp1D-proLYS2-GFP-PKCdelta(C1a+C1b)-NAT
FEATURES             Location/Qualifiers
     source          1..8021
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    102..123
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(138..143,144..161,162..168)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    176..192
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     181..203
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     200..216
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     200..216
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     235..255
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        237..255
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     gene            1232..2351
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     promoter        1232..1575
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1576..2148
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     terminator      2154..2351
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     promoter        3074..3313
                     /gene="S. cerevisiae LYS2"
                     /label=LYS2 promoter
     CDS             3323..4036
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGIDELYK"
     primer_bind     complement(5446..5465)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5447..5465)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5472..5489)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5472..5488)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5481..5503)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5630..6085
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5717..5736)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5927..5948
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6111..6215
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6216..6284,6285..7076)
                     /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     complement(6434..6453)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7247..7835
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7736..7755
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7989..8006
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac
       61 aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact
      121 cattaggcac cccaggcttt acactttatg ctcccggctc gtatgttgtg tggaattgtg
      181 agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa gcgcgcaatt
      241 aaccctcact aaagggaaca aaagctggta ctattagctg aattgccact gctatcgttg
      301 ttagtggcgt tagtgcttgc attcaaagac atggagggcg ttattacgcc ggagctcctc
      361 gacagcagat ctgatgactg gtcaatatat ttttgcattg aggctctgtt tggaattata
      421 ttttgagatg acccatctaa tgtactggta tcaccagatt tcatgtcgtt ttttaaagcg
      481 gctgcttgag tcttagcaat agcgtcacca tctggtgaat cctttgaagg aaccactgac
      541 gaaggtttgg acagtgacga agaggatctt tcctgctttg aattagtcgc gctgggagca
      601 gatgacgagt tggtggagct gggggcagga ttgctggccg tcgtgggtcc tgaatgggtc
      661 cttggctggt ccatctctat tctgaaaacg gaagaggagt agggaatatt actggctgaa
      721 aataagtctt gaatgaacgt atacgcgtat atttctacca atctctcaac actgagtaat
      781 ggtagttata agaaagagac cgagttaggg acagttagag gcggtggaga tattccttat
      841 ggcatgtctg gcgatgataa aacttttcaa acggcagccc cgatctaaaa gagctgacag
      901 ggaaatggtc agaaaaagaa acgtgcaccc gcccgtctgg acgcgccgct cacccgcacg
      961 gcagagacca atcagtaaaa atcaacggtt aacgacatta ctatatatat aatataggaa
     1021 gcatttaata gaacagcatc gtaatatatg tgtactttgc agttatgacg ccagatggca
     1081 gtagtggaag atattcttta ttgaaaaata gcttgtcacc ttacgtacaa tcttgatccg
     1141 gagcttttct ttttttgccg attaagaatt cggtcgaaaa aagaaaagga gagggccaag
     1201 agggagggca ttggtgacta ttgagcacgt ggacatggag gcccagaata ccctccttga
     1261 cagtcttgac gtgcgcagct caggggcatg atgtgactgt cgcccgtaca tttagcccat
     1321 acatccccat gtataatcat ttgcatccat acattttgat ggccgcacgg cgcgaagcaa
     1381 aaattacggc tcctcgctgc agacctgcga gcagggaaac gctcccctca cagacgcgtt
     1441 gaattgtccc cacgccgcgc ccctgtagag aaatataaaa ggttaggatt tgccactgag
     1501 gttcttcttt catatacttc cttttaaaat cttgctagga tacagttctc acatcacatc
     1561 cgaacataaa caaccatggg taccactctt gacgacacgg cttaccggta ccgcaccagt
     1621 gtcccggggg acgccgaggc catcgaggca ctggatgggt ccttcaccac cgacaccgtc
     1681 ttccgcgtca ccgccaccgg ggacggcttc accctgcggg aggtgccggt ggacccgccc
     1741 ctgaccaagg tgttccccga cgacgaatcg gacgacgaat cggacgacgg ggaggacggc
     1801 gacccggact cccggacgtt cgtcgcgtac ggggacgacg gcgacctggc gggcttcgtg
     1861 gtcgtctcgt actccggctg gaaccgccgg ctgaccgtcg aggacatcga ggtcgccccg
     1921 gagcaccggg ggcacggggt cgggcgcgcg ttgatggggc tcgcgacgga gttcgcccgc
     1981 gagcggggcg ccgggcacct ctggctggag gtcaccaacg tcaacgcacc ggcgatccac
     2041 gcgtaccggc ggatggggtt caccctctgc ggcctggaca ccgccctgta cgacggcacc
     2101 gcctcggacg gcgagcaggc gctctacatg agcatgccct gcccctaatc agtactgaca
     2161 ataaaaagat tcttgttttc aagaacttgt catttgtata gtttttttat attgtagttg
     2221 ttctatttta atcaaatgtt agcgtgattt atattttttt tcgcctcgac atcatctgcc
     2281 cagatgcgaa gttaagtgcg cagaaagtaa tatcatgcgt caatcgtatg tgaatgctgg
     2341 tcgctatact gggtaccggg ccccatctgt tgtggcttta tcacgacaga atcttcctca
     2401 attggagcat tcctcttttg aaaaagcctt gaagggtggc tatgtgatcc atgacgtgga
     2461 gaatcctgat attatcctgg tgtcaacagg atcagaagtc tccatttcta tagatgcagc
     2521 caaaaaattg tacgatacta aaaaaatcaa agcaagagtt gtttccctgc cagactttta
     2581 tacttttgac aggcaaagtg aagaatacag attctctgtt ctaccagacg gtgttccgat
     2641 catgtccttt gaagtattgg ctacttcaag ctggggtaag tatgctcatc aatcgttcgg
     2701 actcgacgaa tttagtcgtt caggcaaggg gcctgaaatt tacaaattgt tcgatttcac
     2761 agcggacggt gttgcgtcaa gggctgaaaa gacaatcaat tactacaaag gaaagcagtt
     2821 gctttctcct atgggaagag ctttctaagt ctgaagaagt aaacagttct ttgctatttc
     2881 acacttcctg gttgatggtc acttgctgcc tgaaatatat atatatgtat gacatatgta
     2941 cttgttttct tttttgtgcc tttgttacgt ctatattcat tgaaactgat tattcgattt
     3001 tcttcttgct gaccgcttct agaggcatcg cacagtttta gcgaggaaaa ctcttcaata
     3061 gttttgccag cggaattcca cttgcaatta cataaaaaat tccggcggtt tttcgcgtgt
     3121 gactcaatgt cgaaatacct gcctaatgaa catgaacatc gcccaaatgt atttgaagac
     3181 ccgctgggag aagttcaaga tatataagta acaagcagcc aatagtataa aaaaaaatct
     3241 gagtttatta cctttcctgg aatttcagtg aaaaactgct aattatagag agatatcaca
     3301 gagttactca ctaactagta acatgtctaa aggtgaagaa ttattcactg gtgttgtccc
     3361 aattttggtt gaattagatg gtgatgttaa tggtcacaaa ttttctgtct ccggtgaagg
     3421 tgaaggtgat gctacttacg gtaaattgac cttaaaattt atttgtacta ctggtaaatt
     3481 gccagttcca tggccaacct tagtcactac tttaacttat ggtgttcaat gtttttctag
     3541 atacccagat catatgaaac aacatgactt tttcaagtct gccatgccag aaggttatgt
     3601 tcaagaaaga actatttttt tcaaagatga cggtaactac aagaccagag ctgaagtcaa
     3661 gtttgaaggt gataccttag ttaatagaat cgaattaaaa ggtattgatt ttaaagaaga
     3721 tggtaacatt ttaggtcaca aattggaata caactataac tctcacaatg tttacatcat
     3781 ggctgacaaa caaaagaatg gtatcaaagt taacttcaaa attagacaca acattgaaga
     3841 tggttctgtt caattagctg accattatca acaaaatact ccaattggtg atggtccagt
     3901 cttgttacca gacaaccatt acttatccac tcaatctgcc ttatccaaag atccaaacga
     3961 aaagagagac cacatggtct tgttagaatt tgttactgct gctggtatta cccatggtat
     4021 tgatgaattg tacaaaggag ccggaacaag tgacatgaag cacgagttta tcgccacctt
     4081 cttcgggcag cccaccttct gttctgtgtg caaagagttt gtctggggcc tcaacaagca
     4141 aggctacaaa tgcaggcaat gcaacgctgc catccacaag aaatgcattg acaagattat
     4201 cggccgctgc actggcactg ccaccaatag ccgggacacc atcttccaga aagaacgctt
     4261 caacatcgac atgcctcacc gattcaaggt ttataactac atgagcccca ccttctgtga
     4321 ccactgtggc agtttgctct ggggactggt gaagcaggga ttaaagtgtg aagattgtgg
     4381 catgaatgtg caccacaaat gccgggagaa ggtggccaac ctgtgttagc cgcggtgggt
     4441 tattactgag tagtatttat ttaagtattg tttgtgcact tgcctgcagg ccttttgaaa
     4501 agcaagcata aaagatctaa acataaaatc tgtaaaataa caagatgtaa agataatgct
     4561 aaatcatttg gctttttgat tgattgtaca ggaaaatata catcgcaggg ggttgacttt
     4621 taccatttca ccgcaatgga atcaaacttg ttgaagagaa tgttcacagg cgcatacgct
     4681 acaatgaccc gattcttgct agccttttct cggtcttgca aacaaccgcc ggcagcttag
     4741 tatataaata cacatgtaca tacctctctc cgtatcctcg taatcatttt cttgtattta
     4801 tcgtcttttc gctgtaaaaa ctttatcaca cttatctcaa atacacttat taaccgcttt
     4861 tactattatc ttctacgctg acagtaatat caaacagtga cacatattaa acacagtggt
     4921 ttctttgcat aaacaccatc agcctcaagt cgtcaagtaa agatttcgtg ttcatgcaga
     4981 tagataacaa tctatatgtt gataattagc gttgcctcat caatgcgaga tccgtttaac
     5041 cggaccctag tgcacttacc ccacgttcgg tccactgtgt gccgaacatg ctccttcact
     5101 attttaacat gtggaattct tgaaagaatg aaatcgccat gccaagccat cacacggtct
     5161 tttatgcaat tgattgaccg cctgcaacac ataggcagta aaatttttac tgaaacgtat
     5221 ataatcatca taagcgacaa gtgaggcaac acctttgtta ccacattgac aaccccaggt
     5281 attcatactt cctattagcg gaatcaggag tgcaaaaaga gaaaataaaa gtaaaaaggt
     5341 agggcaacac atagtatgaa tacaaacgtt ccaatattca gttctccggt cagagattta
     5401 ccaaggtctt tcgaacaaaa acatttagcg gttgtagaca attcgcccta tagtgagtcg
     5461 tattacaatt cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc
     5521 caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc
     5581 cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggga cgcgccctgt
     5641 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     5701 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     5761 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     5821 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     5881 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     5941 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     6001 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
     6061 aacaaaatat taacgcttac aatttaggtg gcacttttcg gggaaatgtg cgcggaaccc
     6121 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     6181 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     6241 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     6301 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     6361 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     6421 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     6481 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     6541 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     6601 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     6661 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     6721 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     6781 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     6841 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     6901 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     6961 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     7021 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     7081 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     7141 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     7201 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     7261 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     7321 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     7381 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     7441 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     7501 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     7561 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     7621 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     7681 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     7741 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     7801 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     7861 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     7921 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     7981 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg c
//
