LOCUS       sequence-370826-        8187 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A plasmid for in vivo gene amplification (HamAmp) at SEC23 locus (9 
            copy).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8187)
  TITLE     Vickers Lab plasmids
REFERENCE   2  (bases 1 to 8187)
  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"
COMMENT     Sequence Label: pILGFP5EC4
FEATURES             Location/Qualifiers
     source          1..8187
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(1059..1075)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1059..1075)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1072..1094)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1083..1099
                     /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)."
     promoter        complement(join(1107..1113,1114..1131,1132..1137))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1152..1173
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1290..1307)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1461..2049)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1541..1560)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2220..3011,3012..3080))
                     /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     2843..2862
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3081..3185)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3253..3271
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3309..3331)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3431..3450
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3644..3666
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3658..3675
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3659..3675
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             4632..5660
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             4632..5660
                     /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      5666..5863
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    complement(6610..6643)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        6854..7267
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     CDS             7290..8000
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
ORIGIN
        1 ttacccggga atctcggtcg taatgatttg atatgaaggc atgcccggat atgaaaatgg
       61 aatgctagct caatgggatc taagatcacc ggaatgccca gttggtgagt ttttaattaa
      121 tgaaggcacc ccaataaaca atgtctactt tgcagccgga gccttatttg tatcttctgg
      181 attcgacacc agtattaagc ttgacattat ttcagaccca gaaagtgaac ggcccgctat
      241 tgaatttgag acgcccacct ttttagtttc caatgacgat gaggtcagcc aattctgtta
      301 cgtttctgat gatgaatcaa atggggaggt gcttgaagtt ggaaaaaaca acttctgcgc
      361 tttgtataac ttaagtaatc cttaacctct ttcgtactag taactttttc aaaaaagttc
      421 acttcgctgc atgagcttgg cgtaatgcgc tttgagggtg ataatgcatt ttatttccat
      481 atgaaatatg tctaaaatag aacaattcac taagaacaag aaggccaaac taagcagaat
      541 atggacttcg agactaatga agacatcaac ggggtccgtt tcacctggaa cgtgtttccc
      601 tctactagaa gcgacgctaa cagcaacgtc gtccctgtgg gctgtttgta cacacctttg
      661 aaggagtatg acgaattgaa cgttgcccca tacaatccag ttgtgtgttc cggtccacac
      721 tgtaagtcga ttctaaatcc ttactgtgtg attgatccgc gcaattcgtc ctggagttgt
      781 cccatctgta actccaggaa ccatttacct ccacagtaca ccaacttgtc acaggagaat
      841 atgccgctag agttacagtc caccaccatt gagtacatca ccaataagcc cgttaccgtt
      901 cctcccattt tcttctttgt tgtggatttg acttcggaaa ccgagaacct cgattcgcta
      961 aaggagtcta tcatcacttc tttgtctcta ttacctccca acgcgctgat tgggttgatc
     1021 acttacggaa acgtttaaac aagcttggcg taatcatggt catagctgtt tcctgtgtga
     1081 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     1141 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     1201 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     1261 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     1321 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     1381 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     1441 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     1501 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     1561 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     1621 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     1681 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     1741 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     1801 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     1861 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
     1921 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     1981 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     2041 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     2101 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     2161 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     2221 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     2281 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     2341 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     2401 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     2461 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     2521 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     2581 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     2641 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     2701 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     2761 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     2821 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     2881 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     2941 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     3001 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     3061 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     3121 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     3181 ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca
     3241 ttaacctata aaaataggcg tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac
     3301 ggtgaaaacc tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat
     3361 gccgggagca gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg
     3421 cttaactatg cggcatcaga gcagattgta ctgagagtgc accatatgcg gtgtgaaata
     3481 ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc attcgccatt caggctgcgc
     3541 aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg
     3601 ggatgtgctg caaggcgatt aagttgggta acgccagggt tttcccagtc acgacgttgt
     3661 aaaacgacgg ccagtgaatt cagtttaaac tcttctgctt cgttcagctg caggttaaag
     3721 tagttgtccg tggacacgag cgtacctcta tattcggtgc tgttgaactt aagcttgacg
     3781 cctacacgat ggttgaccag gcctttgagg aacggcttcg ggttcaccgg ctgcatcgcg
     3841 ctgatatcac tgctctcgct cattgtcttg ttgtgttgtg acgtagtgtg tggtgtggta
     3901 tagtacgcgg tgctgtgtgt gctgtgctgt ttttgcgttg ctcatagaac cgttgtttct
     3961 tgctcttacc atctcatgtt catatatgtt ggggaaaaga ggtggaacaa gaaaagttac
     4021 ccggacggca gagcggttgt tttgcctgct ttgccataac acaggatggg atcacacacc
     4081 ccccaagcca tgctacaact tggaagagtg caattcaagt tggttgtcct cagtagtttg
     4141 agagattgcg ctaattttgt tttatcaccg ttatttactg tcagttgaga tttgtcgctt
     4201 gcgacgacta gcccgccgcc tctcgtttgt tacaaaggag gaaagttcta gcaagaataa
     4261 aagaaaaaca cgtttgaaaa aagaaaaaaa gatacgggcc cgaataccct ccttgacagt
     4321 cttgacgtgc gcagctcagg ggcatgatgt gactgtcgcc cgtacattta gcccatacat
     4381 ccccatgtat aatcatttgc atccatacat tttgatggcc gcacggcgcg aagcaaaaat
     4441 tacggctcct cgctgcggac ctgcgagcag ggaaacgctc ccctcacaga cgcgttgaat
     4501 tgtccccacg ccgcgcccct gtagagaaat ataaaaggtt aggatttgcc actgaggttc
     4561 ttctttcata tacttccttt taaaatcttg ctaggataca gttctcacat cacatccgaa
     4621 cataaacaac catgggtaaa aagcctgaac tcaccgcgac gtctgtcgag aagtttctga
     4681 tcgaaaagtt cgacagcgtc tccgacctga tgcagctctc ggagggcgaa gaatctcgtg
     4741 ctttcagctt cgatgtagga gggcgtggat atgtcctgcg ggtaaatagc tgcgccgatg
     4801 gtttctacaa agatcgttat gtttatcggc actttgcatc ggccgcgctc ccgattccgg
     4861 aagtgcttga cattggggaa ttcagcgaga gcctgaccta ttgcatctcc cgccgtgcac
     4921 agggtgtcac gttgcaagac ctgcctgaaa ccgaactgcc cgctgttctg cagccggtcg
     4981 cggaggccat ggatgcgatc gctgcggccg atcttagcca gacgagcggg ttcggcccat
     5041 tcggaccgca aggaatcggt caatacacta catggcgtga tttcatatgc gcgattgctg
     5101 atccccatgt gtatcactgg caaactgtga tggacgacac cgtcagtgcg tccgtcgcgc
     5161 aggctctcga tgagctgatg ctttgggccg aggactgccc cgaagtccgg cacctcgtgc
     5221 acgcggattt cggctccaac aatgtcctga cggacaatgg ccgcataaca gcggtcattg
     5281 actggagcga ggcgatgttc ggggattccc aatacgaggt cgccaacatc ttcttctgga
     5341 ggccgtggtt ggcttgtatg gagcagcaga cgcgctactt cgagcggagg catccggagc
     5401 ttgcaggatc gccgcggctc cgggcgtata tgctccgcat tggtcttgac caactctatc
     5461 agagcttggt tgacggcaat ttcgatgatg cagcttgggc gcagggtcga tgcgacgcaa
     5521 tcgtccgatc cggagccggg actgtcgggc gtacacaaat cgcccgcaga agcgcggccg
     5581 tctggaccga tggctgtgta gaagtactcg ccgatagtgg aaaccgacgc cccagcactc
     5641 gtccgagggc aaaggaataa tcagtactga caataaaaag attcttgttt tcaagaactt
     5701 gtcatttgta tagttttttt atattgtagt tgttctattt taatcaaatg ttagcgtgat
     5761 ttatattttt tttcgcctcg acatcatctg cccagatgcg aagttaagtg cgcagaaagt
     5821 aatatcatgc gtcaatcgta tgtgaatgct ggtcgctata ctgctgtcga ttcgatacta
     5881 acggcggccg cgagcaacgg ctttcttttg ttttatttga tcgttattag gctttttgta
     5941 taatataagg cctgttcagt ccattgcgcg tgcaagaatt ttcctttgga cctgaacgaa
     6001 aagagtagag gattttatat aatatattct ccttcttttt tgtactgcgt ggcctttact
     6061 acgaatatct gtatctaatt tcttttctaa cttttttatt accattacta ctactactat
     6121 tattattatt attattatta ttattattgt gtatgactat aatgctttac tgcctccaat
     6181 tatgcttttt ttattatttt ttttggctga aactacagta tagtaggtat gtatatatct
     6241 aaaactcaaa aagaggcatg tctaatgtaa taccatcaaa aacaatgaaa ttatttaatg
     6301 gagatagcct tttacctaga atgttaatga catcctgcgc tacagcaccc cctataatgg
     6361 ccgcgacagg agcaaactcg atacctttct gtttgataaa ctgttgtatg taatcgtcct
     6421 ttaccactgt tgctggtaca cccagatttt cacaccatac ggccgcatct cttttcattt
     6481 gtatttaaat ccatttcaaa ttttatgttt agttcgagat cctcagtttt cggcgcatag
     6541 gaaccacgta cataataact aaacataaat ctataataaa taaaaaacaa cgatgggagc
     6601 tcgagcctaa taacttcgta tagcatacat tatacgaagt tatattaagg gttgctcgac
     6661 cagaaagcga ccacccaact ttggctgata atagcgtata aacaatgcat actttgtacg
     6721 ttcaaaatac aatgcagtag atatatttat gcatattaca tataatacat atcacatagg
     6781 aagcaacagg cgcgttggac ttttaatttt cgaggaccgc gaatccttac atcacaccca
     6841 atcccccaca agtgatcccc cacacaccat agcttcaaaa tgtttctact ccttttttac
     6901 tcttccagat tttctcggac tccgcgcatc gccgtaccac ttcaaaacac ccaagcacag
     6961 catactaaat ttcccctctt tcttcctcta gggtgtcgtt aattacccgt actaaaggtt
     7021 tggaaaagaa aaaagagacc gcctcgtttc tttttcttcg tcgaaaaagg caataaaaat
     7081 ttttatcacg tttctttttc ttgaaaattt ttttttttga tttttttctc tttcgatgac
     7141 ctcccattga tatttaagtt aataaacggt cttcaatttc tcaagtttca gtttcatttt
     7201 tcttgttcta ttacaacttt ttttacttct tgctcattag aaagaaagca tagcaatcta
     7261 atctaagttt taattacaaa atgggatcct ctaaaggtga agaattattc actggtgttg
     7321 tcccaatttt ggttgaatta gatggtgatg ttaatggtca caaattttct gtctccggtg
     7381 aaggtgaagg tgatgctact tacggtaaat tgaccttaaa atttatttgt actactggta
     7441 aattgccagt tccatggcca accttagtca ctactttcgg ttatggtgtt caatgttttg
     7501 cgagataccc agatcatatg aaacaacatg actttttcaa gtctgccatg ccagaaggtt
     7561 atgttcaaga aagaactatt tttttcaaag atgacggtaa ctacaagacc agagctgaag
     7621 tcaagtttga aggtgatacc ttagttaata gaatcgaatt aaaaggtatt gattttaaag
     7681 aagatggtaa cattttaggt cacaaattgg aatacaacta taactctcac aatgtttaca
     7741 tcatggctga caaacaaaag aatggtatca aagttaactt caaaattaga cacaacattg
     7801 aagatggttc tgttcaatta gctgaccatt atcaacaaaa tactccaatt ggtgatggtc
     7861 cagtcttgtt accagacaac cattacttat ccactcaatc tgccttatcc aaagatccaa
     7921 acgaaaagag agaccacatg gtcttgttag aatttgttac tgctgctggt attacccatg
     7981 gtatggatga attgtacaaa agatcttaaa ctagtaggac tatttgcaaa gggaagggat
     8041 gctaaggtag agggtgaacg ttacagaaaa gcaggctggg aagcatattt gagaagatgc
     8101 ggccagcaaa actaaaaaac tgtattataa gtaaatgcat gtatactaaa ctcacaaatt
     8161 agagcttcaa tttaattata tcagtta
//
