LOCUS       sequence-370818-        7958 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A plasmid for in vivo gene amplification (HamAmp) at RPL25 locus 
            (~12 copy).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7958)
  TITLE     Vickers Lab plasmids
REFERENCE   2  (bases 1 to 7958)
  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: pILGFP3AG4
FEATURES             Location/Qualifiers
     source          1..7958
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(38..54)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(38..54)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(51..73)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    62..78
                     /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(86..92,93..110,111..116))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    131..152
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(269..286)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(440..1028)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(520..539)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1199..1990,1991..2059))
                     /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     1822..1841
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2060..2164)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2232..2250
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2288..2310)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2410..2429
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2623..2645
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2637..2654
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2638..2654
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(5326..5359)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        5570..5983
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     CDS             6006..6716
                     /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 caaggaacat agtttaaaca agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
       61 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
      121 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
      181 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
      241 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
      301 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
      361 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
      421 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
      481 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
      541 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
      601 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
      661 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
      721 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
      781 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
      841 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
      901 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
      961 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     1021 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     1081 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     1141 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     1201 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     1261 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     1321 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     1381 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     1441 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     1501 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     1561 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     1621 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     1681 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     1741 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     1801 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     1861 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     1921 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     1981 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     2041 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     2101 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     2161 cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga aaccattatt atcatgacat
     2221 taacctataa aaataggcgt atcacgaggc cctttcgtct cgcgcgtttc ggtgatgacg
     2281 gtgaaaacct ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg
     2341 ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggctggc
     2401 ttaactatgc ggcatcagag cagattgtac tgagagtgca ccatatgcgg tgtgaaatac
     2461 cgcacagatg cgtaaggaga aaataccgca tcaggcgcca ttcgccattc aggctgcgca
     2521 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg gcgaaagggg
     2581 gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca cgacgttgta
     2641 aaacgacggc cagtgaattc agtttaaaca tgtactaatc agtctaacac cccggaggag
     2701 gcttatcatt acatattctt gtctcggcgt tctttaagtt tcttatcttg tatgcccgat
     2761 atagcaacct tgttggtacc aatctaacgg tttccgtact ttgcaatgaa gagatgagga
     2821 ggcatgggtc acttatttaa tatgtacggg tgtttacatg gagttgcttt ctttttttgt
     2881 ctcagcagtc attgtgcgcc aaaaaaagag aaaaccgtga gccgaagtcc acgctctgga
     2941 gttaggctct cccattacgg agagaagcat ttcctcagcc tgggagcccc gttggaacag
     3001 tcaggctaaa ctgggccttc ctacccactg cttgctgttt ctcactggac gcacaagggg
     3061 attttctttc taccttcggc ttgcctcact gcgttggggc ttcccaatgc aacttcgttc
     3121 gtatgcatac aatcttttag atattatctt ttaaaattat tttaaaacaa ttttaaatgt
     3181 atctcatatg cttttcttct gctgttgaaa aggctaaaca aagaagatca ataagataaa
     3241 gcggccgcaa aatgtctaag aatatcgttg tcctaccggg tgatcacgtc ggtaaagaag
     3301 ttactgacga agctattaag gtcttgaatg ccattgctga agtccgtcca gaaattaagt
     3361 tcaatttcca acatcacttg atcgggggtg ctgccatcga tgccactggc actcctttac
     3421 cagatgaagc tctagaagcc tctaagaaag ccgatgctgt cttactaggt gctgttggtg
     3481 gtccaaaatg gggtacgggc gcagttagac cagaacaagg tctattgaag atcagaaagg
     3541 aattgggtct atacgccaac ttgagaccat gtaactttgc ttctgattct ttactagatc
     3601 tttctccttt gaagcctgaa tatgcaaagg gtaccgattt cgtcgtcgtt agagaattgg
     3661 ttggtggtat ctactttggt gaaagaaaag aagatgaagg tgacggagtt gcttgggact
     3721 ctgagaaata cagtgttcct gaagttcaaa gaattacaag aatggctgct ttcttggcat
     3781 tgcaacaaaa cccaccatta ccaatctggt ctcttgacaa ggctaacgtg cttgcctctt
     3841 ccagattgtg gagaaagact gttgaagaaa ccatcaagac tgagttccca caattaactg
     3901 ttcagcacca attgatcgac tctgctgcta tgattttggt taaatcacca actaagctaa
     3961 acggtgttgt tattaccaac aacatgtttg gtgatattat ctccgatgaa gcctctgtta
     4021 ttccaggttc tttgggttta ttaccttctg catctctagc ttccctacct gacactaaca
     4081 aggcattcgg tttgtacgaa ccatgtcatg gttctgcccc agatttacca gcaaacaagg
     4141 ttaacccaat tgctaccatc ttatctgcag ctatgatgtt gaagttatcc ttggatttgg
     4201 ttgaagaagg tagggctctt gaagaagctg ttagaaatgt cttggatgca ggtgtcagaa
     4261 ccggtgacct tggtggttct aactctacca ctgaggttgg cgatgctatc gccaaggctg
     4321 tcaaggaaat cttggcttaa agagtctttt gtaacgaccc cgtctccacc aacttggtat
     4381 gcttgaaatc tcaaggccat tacacattca gttatgtgaa cgaaaggtct ttatttaacg
     4441 tagcataaac taaataatac aggttccggt tagcctgcgg gcccaaagct agctctaatt
     4501 ggtttaatta ataaatttaa tattattttt aaatttttct ttaaatatac aataaatctt
     4561 tcataacatg ttaaattcat gattaagcgt aaataaagtg tagtggcaga gtgcacgggg
     4621 tttcctgtgc cttacaaagt aggtaccaat ttgcgtattg cagcgagggt tccggttact
     4681 atttataatt acgtgttagt gtactgtgat tttattgagg ctataacaag aaaaggatct
     4741 gtgaaggttt tgagagaatg aggcgaaaac acagggttat tttcatcagc aacgccgctg
     4801 tttcagcggc accctgccct tttcttttcg caagatgaga gtaaagagtt gtacatcagg
     4861 taagaatgtt attatttaaa ttcgaagtga taaattcttt tcatgatgaa tcactcgctt
     4921 atatggggta gaatatatat atatgtgtgt gtgtgtgtgt gtttgtgtat gtagggatgg
     4981 tgcgcgtttg ttgtgtgaca tttgctactc attcttttcc ttttcctacg actggcttaa
     5041 cgggaatatt atcaatttgc tgcattctta tgcttcggtc cgatgctcaa gcttaacttc
     5101 ttcgtgagga aggaaagtgt tgtctcctac ttttttcaaa ttttcgaatt gtatttatat
     5161 ttatttagta cttcttgagt ttacatatcc ttcgtaaaaa tgcaactttt gtcgaaaaac
     5221 acttccaaaa aaaaataata atgaatttat gaagcatact aacgagcgag cacatcgctg
     5281 aggtatcatt acttcatgag ataaattaac tcgagctcga gcctaataac ttcgtatagc
     5341 atacattata cgaagttata ttaagggttg ctcgaccaga aagcgaccac ccaactttgg
     5401 ctgataatag cgtataaaca atgcatactt tgtacgttca aaatacaatg cagtagatat
     5461 atttatgcat attacatata atacatatca cataggaagc aacaggcgcg ttggactttt
     5521 aattttcgag gaccgcgaat ccttacatca cacccaatcc cccacaagtg atcccccaca
     5581 caccatagct tcaaaatgtt tctactcctt ttttactctt ccagattttc tcggactccg
     5641 cgcatcgccg taccacttca aaacacccaa gcacagcata ctaaatttcc cctctttctt
     5701 cctctagggt gtcgttaatt acccgtacta aaggtttgga aaagaaaaaa gagaccgcct
     5761 cgtttctttt tcttcgtcga aaaaggcaat aaaaattttt atcacgtttc tttttcttga
     5821 aaattttttt ttttgatttt tttctctttc gatgacctcc cattgatatt taagttaata
     5881 aacggtcttc aatttctcaa gtttcagttt catttttctt gttctattac aacttttttt
     5941 acttcttgct cattagaaag aaagcatagc aatctaatct aagttttaat tacaaaatgg
     6001 gatcctctaa aggtgaagaa ttattcactg gtgttgtccc aattttggtt gaattagatg
     6061 gtgatgttaa tggtcacaaa ttttctgtct ccggtgaagg tgaaggtgat gctacttacg
     6121 gtaaattgac cttaaaattt atttgtacta ctggtaaatt gccagttcca tggccaacct
     6181 tagtcactac tttcggttat ggtgttcaat gttttgcgag atacccagat catatgaaac
     6241 aacatgactt tttcaagtct gccatgccag aaggttatgt tcaagaaaga actatttttt
     6301 tcaaagatga cggtaactac aagaccagag ctgaagtcaa gtttgaaggt gataccttag
     6361 ttaatagaat cgaattaaaa ggtattgatt ttaaagaaga tggtaacatt ttaggtcaca
     6421 aattggaata caactataac tctcacaatg tttacatcat ggctgacaaa caaaagaatg
     6481 gtatcaaagt taacttcaaa attagacaca acattgaaga tggttctgtt caattagctg
     6541 accattatca acaaaatact ccaattggtg atggtccagt cttgttacca gacaaccatt
     6601 acttatccac tcaatctgcc ttatccaaag atccaaacga aaagagagac cacatggtct
     6661 tgttagaatt tgttactgct gctggtatta cccatggtat ggatgaattg tacaaaagat
     6721 cttaaactag taggactatt tgcaaaggga agggatgcta aggtagaggg tgaacgttac
     6781 agaaaagcag gctgggaagc atatttgaga agatgcggcc agcaaaacta aaaaactgta
     6841 ttataagtaa atgcatgtat actaaactca caaattagag cttcaattta attatatcag
     6901 ttattacccg ggaatctcgg tcgtaatgat ttgcatgcga aattcaaaac tctccagaca
     6961 aagcctgcca tttggccaag caagcttttg acgacgctat tgctgagttg gacactctgt
     7021 ctgaagaatc atacaaagat agcacactta tcatgcaact gctaagggac aatttaacct
     7081 tatggacttc agacatgtcc gagtccggtc aagctgaaga ccaacaacaa caacaacaac
     7141 atcagcaaca gcagccacct gctgccgccg aaggtgaagc accaaagtaa gtattccgat
     7201 aaatctaaag aaaaattact aaaaaaaaga aaaaaaaaag aacgggggta taataatttg
     7261 tagttcatta ttgcaattat atatctatat ctatatatgt atataacatt aacatgtgca
     7321 cgtacacacg taatcgcgcg tgtacatgtc tatatgtgtt acttgaacta tactgttttg
     7381 acgtgtatgt ttgtttatct ctcttctgat tcctccaccc cttccttact caaccgggta
     7441 aatgtcgcat catgactccc gacaataatc ccctctggta tagcgagaag caacttcagc
     7501 ttcttaacgg caagaacttt tttatgtttg tcgcacctgt agcttcacaa aagttggata
     7561 cagcaataag aaaggaaacc acatttgtgc caatggctcc atctggtatg tgaactgcaa
     7621 tattaatagc acgagaaaat tgagaggaag atagatggga actagtagag ttgatattga
     7681 tgagatacga aaaccacacg taaataaact atccgacgac aagaatagtg cattaaggac
     7741 ttgattaaga tatggtgagc aacgtaatta tcgggctcaa cagtttatta gcaatcgttt
     7801 tgatagaagc gttgatgctg tggaagttgt tctttttact accaacagcc attaacaaat
     7861 ccattagagt gttcgttcgt ttttctgctc agattcaaga aaattattcc atcctcatta
     7921 tacttttttc ttctatttcg tgctccacgt cgaggtat
//
