LOCUS       sequence-370821-        8095 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A plasmid for in vivo gene amplification (HamAmp) at SEC23 locus (1 
            copy reference).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8095)
  TITLE     Vickers Lab plasmids
REFERENCE   2  (bases 1 to 8095)
  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: pILGFP5EG3
FEATURES             Location/Qualifiers
     source          1..8095
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(964..980)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(964..980)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(977..999)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    988..1004
                     /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(1012..1018,1019..1036,1037..1042))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1057..1078
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1195..1212)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1366..1954)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1446..1465)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2125..2916,2917..2985))
                     /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     2748..2767
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2986..3090)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3158..3176
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3214..3236)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3336..3355
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3549..3571
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3563..3580
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3564..3580
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             4537..5565
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             4537..5565
                     /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      5571..5768
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    complement(6515..6548)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        6759..7172
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     CDS             7195..7905
                     /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 cccgggaatc tcggtcgtaa tgatttgata tgaaggcatg ctgcgatact gccgtagcgg
       61 gccttcgtat agctcggccg agctcgtaca aaaggcaagc agtgtatcgg acagagctga
      121 tataacacaa tacgctcgta gtcgatgcat gccgtggctg ctctcggtcg ggtataagtc
      181 ttagacaata gtcttacctc gcatgtataa taaatctttt gtatttaatc tattatatgt
      241 ttctatgctt ttttttccta ttgttgtttg cttttccttt tccttatttc tttctagctt
      301 ctaattttct ttcttttttt ttttttttca ttgaaaatta tatatatata tatatatcag
      361 aacaattgtc cagtattgaa caatacaggt tatttcgaac aattgaaaaa aaaaaatcac
      421 agaaaaacat atcgagaaaa gggtcatgga cttcgagact aatgaagaca tcaacggggt
      481 ccgtttcacc tggaacgtgt ttccctctac tagaagcgac gctaacagca acgtcgtccc
      541 tgtgggctgt ttgtacacac ctttgaagga gtatgacgaa ttgaacgttg ccccatacaa
      601 tccagttgtg tgttccggtc cacactgtaa gtcgattcta aatccttact gtgtgattga
      661 tccgcgcaat tcgtcctgga gttgtcccat ctgtaactcc aggaaccatt tacctccaca
      721 gtacaccaac ttgtcacagg agaatatgcc gctagagtta cagtccacca ccattgagta
      781 catcaccaat aagcccgtta ccgttcctcc cattttcttc tttgttgtgg atttgacttc
      841 ggaaaccgag aacctcgatt cgctaaagga gtctatcatc acttctttgt ctctattacc
      901 tcccaacgcg ctgattgggt tgatcactta cggaaacgtt taaacaagct tggcgtaatc
      961 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     1021 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     1081 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     1141 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     1201 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     1261 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     1321 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     1381 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     1441 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     1501 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     1561 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     1621 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     1681 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     1741 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     1801 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     1861 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     1921 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     1981 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     2041 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     2101 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     2161 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     2221 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     2281 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     2341 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     2401 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     2461 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     2521 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     2581 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     2641 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     2701 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     2761 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     2821 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     2881 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     2941 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     3001 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     3061 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
     3121 ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt
     3181 tcgtctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac
     3241 ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc
     3301 gggtgttggc gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag
     3361 agtgcaccat atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag
     3421 gcgccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc
     3481 gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc
     3541 agggttttcc cagtcacgac gttgtaaaac gacggccagt gaattcagtt taaactcttc
     3601 tgcttcgttc agctgcaggt taaagtagtt gtccgtggac acgagcgtac ctctatattc
     3661 ggtgctgttg aacttaagct tgacgcctac acgatggttg accaggcctt tgaggaacgg
     3721 cttcgggttc accggctgca tcgcgctgat atcactgctc tcgctcattg tcttgttgtg
     3781 ttgtgacgta gtgtgtggtg tggtatagta cgcggtgctg tgtgtgctgt gctgtttttg
     3841 cgttgctcat agaaccgttg tttcttgctc ttaccatctc atgttcatat atgttgggga
     3901 aaagaggtgg aacaagaaaa gttacccgga cggcagagcg gttgttttgc ctgctttgcc
     3961 ataacacagg atgggatcac acacccccca agccatgcta caacttggaa gagtgcaatt
     4021 caagttggtt gtcctcagta gtttgagaga ttgcgctaat tttgttttat caccgttatt
     4081 tactgtcagt tgagatttgt cgcttgcgac gactagcccg ccgcctctcg tttgttacaa
     4141 aggaggaaag ttctagcaag aataaaagaa aaacacgttt gaaaaaagaa aaaaagatac
     4201 gggcccgaat accctccttg acagtcttga cgtgcgcagc tcaggggcat gatgtgactg
     4261 tcgcccgtac atttagccca tacatcccca tgtataatca tttgcatcca tacattttga
     4321 tggccgcacg gcgcgaagca aaaattacgg ctcctcgctg cggacctgcg agcagggaaa
     4381 cgctcccctc acagacgcgt tgaattgtcc ccacgccgcg cccctgtaga gaaatataaa
     4441 aggttaggat ttgccactga ggttcttctt tcatatactt ccttttaaaa tcttgctagg
     4501 atacagttct cacatcacat ccgaacataa acaaccatgg gtaaaaagcc tgaactcacc
     4561 gcgacgtctg tcgagaagtt tctgatcgaa aagttcgaca gcgtctccga cctgatgcag
     4621 ctctcggagg gcgaagaatc tcgtgctttc agcttcgatg taggagggcg tggatatgtc
     4681 ctgcgggtaa atagctgcgc cgatggtttc tacaaagatc gttatgttta tcggcacttt
     4741 gcatcggccg cgctcccgat tccggaagtg cttgacattg gggaattcag cgagagcctg
     4801 acctattgca tctcccgccg tgcacagggt gtcacgttgc aagacctgcc tgaaaccgaa
     4861 ctgcccgctg ttctgcagcc ggtcgcggag gccatggatg cgatcgctgc ggccgatctt
     4921 agccagacga gcgggttcgg cccattcgga ccgcaaggaa tcggtcaata cactacatgg
     4981 cgtgatttca tatgcgcgat tgctgatccc catgtgtatc actggcaaac tgtgatggac
     5041 gacaccgtca gtgcgtccgt cgcgcaggct ctcgatgagc tgatgctttg ggccgaggac
     5101 tgccccgaag tccggcacct cgtgcacgcg gatttcggct ccaacaatgt cctgacggac
     5161 aatggccgca taacagcggt cattgactgg agcgaggcga tgttcgggga ttcccaatac
     5221 gaggtcgcca acatcttctt ctggaggccg tggttggctt gtatggagca gcagacgcgc
     5281 tacttcgagc ggaggcatcc ggagcttgca ggatcgccgc ggctccgggc gtatatgctc
     5341 cgcattggtc ttgaccaact ctatcagagc ttggttgacg gcaatttcga tgatgcagct
     5401 tgggcgcagg gtcgatgcga cgcaatcgtc cgatccggag ccgggactgt cgggcgtaca
     5461 caaatcgccc gcagaagcgc ggccgtctgg accgatggct gtgtagaagt actcgccgat
     5521 agtggaaacc gacgccccag cactcgtccg agggcaaagg aataatcagt actgacaata
     5581 aaaagattct tgttttcaag aacttgtcat ttgtatagtt tttttatatt gtagttgttc
     5641 tattttaatc aaatgttagc gtgatttata ttttttttcg cctcgacatc atctgcccag
     5701 atgcgaagtt aagtgcgcag aaagtaatat catgcgtcaa tcgtatgtga atgctggtcg
     5761 ctatactgct gtcgattcga tactaacggc ggccgcgagc aacggctttc ttttgtttta
     5821 tttgatcgtt attaggcttt ttgtataata taaggcctgt tcagtccatt gcgcgtgcaa
     5881 gaattttcct ttggacctga acgaaaagag tagaggattt tatataatat attctccttc
     5941 ttttttgtac tgcgtggcct ttactacgaa tatctgtatc taatttcttt tctaactttt
     6001 ttattaccat tactactact actattatta ttattattat tattattatt attgtgtatg
     6061 actataatgc tttactgcct ccaattatgc tttttttatt attttttttg gctgaaacta
     6121 cagtatagta ggtatgtata tatctaaaac tcaaaaagag gcatgtctaa tgtaatacca
     6181 tcaaaaacaa tgaaattatt taatggagat agccttttac ctagaatgtt aatgacatcc
     6241 tgcgctacag caccccctat aatggccgcg acaggagcaa actcgatacc tttctgtttg
     6301 ataaactgtt gtatgtaatc gtcctttacc actgttgctg gtacacccag attttcacac
     6361 catacggccg catctctttt catttgtatt taaatccatt tcaaatttta tgtttagttc
     6421 gagatcctca gttttcggcg cataggaacc acgtacataa taactaaaca taaatctata
     6481 ataaataaaa aacaacgatg ggagctcgag cctaataact tcgtatagca tacattatac
     6541 gaagttatat taagggttgc tcgaccagaa agcgaccacc caactttggc tgataatagc
     6601 gtataaacaa tgcatacttt gtacgttcaa aatacaatgc agtagatata tttatgcata
     6661 ttacatataa tacatatcac ataggaagca acaggcgcgt tggactttta attttcgagg
     6721 accgcgaatc cttacatcac acccaatccc ccacaagtga tcccccacac accatagctt
     6781 caaaatgttt ctactccttt tttactcttc cagattttct cggactccgc gcatcgccgt
     6841 accacttcaa aacacccaag cacagcatac taaatttccc ctctttcttc ctctagggtg
     6901 tcgttaatta cccgtactaa aggtttggaa aagaaaaaag agaccgcctc gtttcttttt
     6961 cttcgtcgaa aaaggcaata aaaattttta tcacgtttct ttttcttgaa aatttttttt
     7021 tttgattttt ttctctttcg atgacctccc attgatattt aagttaataa acggtcttca
     7081 atttctcaag tttcagtttc atttttcttg ttctattaca acttttttta cttcttgctc
     7141 attagaaaga aagcatagca atctaatcta agttttaatt acaaaatggg atcctctaaa
     7201 ggtgaagaat tattcactgg tgttgtccca attttggttg aattagatgg tgatgttaat
     7261 ggtcacaaat tttctgtctc cggtgaaggt gaaggtgatg ctacttacgg taaattgacc
     7321 ttaaaattta tttgtactac tggtaaattg ccagttccat ggccaacctt agtcactact
     7381 ttcggttatg gtgttcaatg ttttgcgaga tacccagatc atatgaaaca acatgacttt
     7441 ttcaagtctg ccatgccaga aggttatgtt caagaaagaa ctattttttt caaagatgac
     7501 ggtaactaca agaccagagc tgaagtcaag tttgaaggtg ataccttagt taatagaatc
     7561 gaattaaaag gtattgattt taaagaagat ggtaacattt taggtcacaa attggaatac
     7621 aactataact ctcacaatgt ttacatcatg gctgacaaac aaaagaatgg tatcaaagtt
     7681 aacttcaaaa ttagacacaa cattgaagat ggttctgttc aattagctga ccattatcaa
     7741 caaaatactc caattggtga tggtccagtc ttgttaccag acaaccatta cttatccact
     7801 caatctgcct tatccaaaga tccaaacgaa aagagagacc acatggtctt gttagaattt
     7861 gttactgctg ctggtattac ccatggtatg gatgaattgt acaaaagatc ttaaactagt
     7921 aggactattt gcaaagggaa gggatgctaa ggtagagggt gaacgttaca gaaaagcagg
     7981 ctgggaagca tatttgagaa gatgcggcca gcaaaactaa aaaactgtat tataagtaaa
     8041 tgcatgtata ctaaactcac aaattagagc ttcaatttaa ttatatcagt tatta
//
