LOCUS       sequence-370828-        8196 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A plasmid for in vivo gene amplification (HamAmp) at SEC23 locus (13
            copy).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8196)
  TITLE     Vickers Lab plasmids
REFERENCE   2  (bases 1 to 8196)
  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: pILGFP5EF3
FEATURES             Location/Qualifiers
     source          1..8196
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(1069..1085)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1069..1085)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1082..1104)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1093..1109
                     /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(1117..1123,1124..1141,1142..1147))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1162..1183
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1300..1317)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1471..2059)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1551..1570)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2230..3021,3022..3090))
                     /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     2853..2872
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3091..3195)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3263..3281
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3319..3341)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3441..3460
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3654..3676
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3668..3685
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3669..3685
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             4642..5670
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             4642..5670
                     /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      5676..5873
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    complement(6620..6653)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        6864..7277
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     CDS             7300..8010
                     /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 attacccggg aatctcggtc gtaatgattt gatatgaagg catgcccgga tatgaaaatg
       61 gaatgctagc tcaatgggat ctaagatcac cggaatgccc agttggtgag tttttaatta
      121 atgaaggcac cccaataaac aatgtctact ttgcagccgg agccttattt gtatcttctg
      181 gattcgacac cagtattaag cttgacatta tttcagaccc agaaagtgaa cggcccgcta
      241 ttgaatttga gacgcccacc tttttagttt ccaatgacga tgaggtcagc caattctgtt
      301 acgtttctga tgatgaatca aatggggagg tgcttgaagt tggaaaaaac aacttctgcg
      361 ctttgtataa cttaagtaat ccttaacctc tttcgtacta gtaacttttt caaaaaagtt
      421 cacttcgctg catgagcttg gcgtaatgcg ctttgagggt gataatgcat tttatttcca
      481 tatgaaatat gtctaaaata gaacaattca ctaagaacaa gaaggccaaa ctaagcagaa
      541 tatgggagga ggagacttcg agactaatga agacatcaac ggggtccgtt tcacctggaa
      601 cgtgtttccc tctactagaa gcgacgctaa cagcaacgtc gtccctgtgg gctgtttgta
      661 cacacctttg aaggagtatg acgaattgaa cgttgcccca tacaatccag ttgtgtgttc
      721 cggtccacac tgtaagtcga ttctaaatcc ttactgtgtg attgatccgc gcaattcgtc
      781 ctggagttgt cccatctgta actccaggaa ccatttacct ccacagtaca ccaacttgtc
      841 acaggagaat atgccgctag agttacagtc caccaccatt gagtacatca ccaataagcc
      901 cgttaccgtt cctcccattt tcttctttgt tgtggatttg acttcggaaa ccgagaacct
      961 cgattcgcta aaggagtcta tcatcacttc tttgtctcta ttacctccca acgcgctgat
     1021 tgggttgatc acttacggaa acgtttaaac aagcttggcg taatcatggt catagctgtt
     1081 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
     1141 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     1201 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     1261 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     1321 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     1381 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     1441 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     1501 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     1561 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     1621 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     1681 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     1741 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     1801 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     1861 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     1921 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     1981 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     2041 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     2101 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     2161 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     2221 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     2281 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     2341 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     2401 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     2461 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     2521 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     2581 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     2641 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     2701 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     2761 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     2821 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     2881 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     2941 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     3001 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     3061 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     3121 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     3181 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccattat
     3241 tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc tcgcgcgttt
     3301 cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca cagcttgtct
     3361 gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg
     3421 tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc accatatgcg
     3481 gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc attcgccatt
     3541 caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct
     3601 ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt tttcccagtc
     3661 acgacgttgt aaaacgacgg ccagtgaatt cagtttaaac tcttctgctt cgttcagctg
     3721 caggttaaag tagttgtccg tggacacgag cgtacctcta tattcggtgc tgttgaactt
     3781 aagcttgacg cctacacgat ggttgaccag gcctttgagg aacggcttcg ggttcaccgg
     3841 ctgcatcgcg ctgatatcac tgctctcgct cattgtcttg ttgtgttgtg acgtagtgtg
     3901 tggtgtggta tagtacgcgg tgctgtgtgt gctgtgctgt ttttgcgttg ctcatagaac
     3961 cgttgtttct tgctcttacc atctcatgtt catatatgtt ggggaaaaga ggtggaacaa
     4021 gaaaagttac ccggacggca gagcggttgt tttgcctgct ttgccataac acaggatggg
     4081 atcacacacc ccccaagcca tgctacaact tggaagagtg caattcaagt tggttgtcct
     4141 cagtagtttg agagattgcg ctaattttgt tttatcaccg ttatttactg tcagttgaga
     4201 tttgtcgctt gcgacgacta gcccgccgcc tctcgtttgt tacaaaggag gaaagttcta
     4261 gcaagaataa aagaaaaaca cgtttgaaaa aagaaaaaaa gatacgggcc cgaataccct
     4321 ccttgacagt cttgacgtgc gcagctcagg ggcatgatgt gactgtcgcc cgtacattta
     4381 gcccatacat ccccatgtat aatcatttgc atccatacat tttgatggcc gcacggcgcg
     4441 aagcaaaaat tacggctcct cgctgcggac ctgcgagcag ggaaacgctc ccctcacaga
     4501 cgcgttgaat tgtccccacg ccgcgcccct gtagagaaat ataaaaggtt aggatttgcc
     4561 actgaggttc ttctttcata tacttccttt taaaatcttg ctaggataca gttctcacat
     4621 cacatccgaa cataaacaac catgggtaaa aagcctgaac tcaccgcgac gtctgtcgag
     4681 aagtttctga tcgaaaagtt cgacagcgtc tccgacctga tgcagctctc ggagggcgaa
     4741 gaatctcgtg ctttcagctt cgatgtagga gggcgtggat atgtcctgcg ggtaaatagc
     4801 tgcgccgatg gtttctacaa agatcgttat gtttatcggc actttgcatc ggccgcgctc
     4861 ccgattccgg aagtgcttga cattggggaa ttcagcgaga gcctgaccta ttgcatctcc
     4921 cgccgtgcac agggtgtcac gttgcaagac ctgcctgaaa ccgaactgcc cgctgttctg
     4981 cagccggtcg cggaggccat ggatgcgatc gctgcggccg atcttagcca gacgagcggg
     5041 ttcggcccat tcggaccgca aggaatcggt caatacacta catggcgtga tttcatatgc
     5101 gcgattgctg atccccatgt gtatcactgg caaactgtga tggacgacac cgtcagtgcg
     5161 tccgtcgcgc aggctctcga tgagctgatg ctttgggccg aggactgccc cgaagtccgg
     5221 cacctcgtgc acgcggattt cggctccaac aatgtcctga cggacaatgg ccgcataaca
     5281 gcggtcattg actggagcga ggcgatgttc ggggattccc aatacgaggt cgccaacatc
     5341 ttcttctgga ggccgtggtt ggcttgtatg gagcagcaga cgcgctactt cgagcggagg
     5401 catccggagc ttgcaggatc gccgcggctc cgggcgtata tgctccgcat tggtcttgac
     5461 caactctatc agagcttggt tgacggcaat ttcgatgatg cagcttgggc gcagggtcga
     5521 tgcgacgcaa tcgtccgatc cggagccggg actgtcgggc gtacacaaat cgcccgcaga
     5581 agcgcggccg tctggaccga tggctgtgta gaagtactcg ccgatagtgg aaaccgacgc
     5641 cccagcactc gtccgagggc aaaggaataa tcagtactga caataaaaag attcttgttt
     5701 tcaagaactt gtcatttgta tagttttttt atattgtagt tgttctattt taatcaaatg
     5761 ttagcgtgat ttatattttt tttcgcctcg acatcatctg cccagatgcg aagttaagtg
     5821 cgcagaaagt aatatcatgc gtcaatcgta tgtgaatgct ggtcgctata ctgctgtcga
     5881 ttcgatacta acggcggccg cgagcaacgg ctttcttttg ttttatttga tcgttattag
     5941 gctttttgta taatataagg cctgttcagt ccattgcgcg tgcaagaatt ttcctttgga
     6001 cctgaacgaa aagagtagag gattttatat aatatattct ccttcttttt tgtactgcgt
     6061 ggcctttact acgaatatct gtatctaatt tcttttctaa cttttttatt accattacta
     6121 ctactactat tattattatt attattatta ttattattgt gtatgactat aatgctttac
     6181 tgcctccaat tatgcttttt ttattatttt ttttggctga aactacagta tagtaggtat
     6241 gtatatatct aaaactcaaa aagaggcatg tctaatgtaa taccatcaaa aacaatgaaa
     6301 ttatttaatg gagatagcct tttacctaga atgttaatga catcctgcgc tacagcaccc
     6361 cctataatgg ccgcgacagg agcaaactcg atacctttct gtttgataaa ctgttgtatg
     6421 taatcgtcct ttaccactgt tgctggtaca cccagatttt cacaccatac ggccgcatct
     6481 cttttcattt gtatttaaat ccatttcaaa ttttatgttt agttcgagat cctcagtttt
     6541 cggcgcatag gaaccacgta cataataact aaacataaat ctataataaa taaaaaacaa
     6601 cgatgggagc tcgagcctaa taacttcgta tagcatacat tatacgaagt tatattaagg
     6661 gttgctcgac cagaaagcga ccacccaact ttggctgata atagcgtata aacaatgcat
     6721 actttgtacg ttcaaaatac aatgcagtag atatatttat gcatattaca tataatacat
     6781 atcacatagg aagcaacagg cgcgttggac ttttaatttt cgaggaccgc gaatccttac
     6841 atcacaccca atcccccaca agtgatcccc cacacaccat agcttcaaaa tgtttctact
     6901 ccttttttac tcttccagat tttctcggac tccgcgcatc gccgtaccac ttcaaaacac
     6961 ccaagcacag catactaaat ttcccctctt tcttcctcta gggtgtcgtt aattacccgt
     7021 actaaaggtt tggaaaagaa aaaagagacc gcctcgtttc tttttcttcg tcgaaaaagg
     7081 caataaaaat ttttatcacg tttctttttc ttgaaaattt ttttttttga tttttttctc
     7141 tttcgatgac ctcccattga tatttaagtt aataaacggt cttcaatttc tcaagtttca
     7201 gtttcatttt tcttgttcta ttacaacttt ttttacttct tgctcattag aaagaaagca
     7261 tagcaatcta atctaagttt taattacaaa atgggatcct ctaaaggtga agaattattc
     7321 actggtgttg tcccaatttt ggttgaatta gatggtgatg ttaatggtca caaattttct
     7381 gtctccggtg aaggtgaagg tgatgctact tacggtaaat tgaccttaaa atttatttgt
     7441 actactggta aattgccagt tccatggcca accttagtca ctactttcgg ttatggtgtt
     7501 caatgttttg cgagataccc agatcatatg aaacaacatg actttttcaa gtctgccatg
     7561 ccagaaggtt atgttcaaga aagaactatt tttttcaaag atgacggtaa ctacaagacc
     7621 agagctgaag tcaagtttga aggtgatacc ttagttaata gaatcgaatt aaaaggtatt
     7681 gattttaaag aagatggtaa cattttaggt cacaaattgg aatacaacta taactctcac
     7741 aatgtttaca tcatggctga caaacaaaag aatggtatca aagttaactt caaaattaga
     7801 cacaacattg aagatggttc tgttcaatta gctgaccatt atcaacaaaa tactccaatt
     7861 ggtgatggtc cagtcttgtt accagacaac cattacttat ccactcaatc tgccttatcc
     7921 aaagatccaa acgaaaagag agaccacatg gtcttgttag aatttgttac tgctgctggt
     7981 attacccatg gtatggatga attgtacaaa agatcttaaa ctagtaggac tatttgcaaa
     8041 gggaagggat gctaaggtag agggtgaacg ttacagaaaa gcaggctggg aagcatattt
     8101 gagaagatgc ggccagcaaa actaaaaaac tgtattataa gtaaatgcat gtatactaaa
     8161 ctcacaaatt agagcttcaa tttaattata tcagtt
//
