LOCUS       sequence-36640-e        7386 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7386)
  AUTHORS   Stowers RS
  TITLE     An efficient method for recombineering GAL4 and QF drivers.
  JOURNAL   Fly (Austin). 2011 Oct 1;5(4).
  PUBMED    21857163
REFERENCE   2  (bases 1 to 7386)
  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"; journalName: "Fly 
            (Austin). 2011 Oct 1;5(4)."
COMMENT     Sequence Label: pGAL4Kan
FEATURES             Location/Qualifiers
     source          1..7386
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        24..128
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(129..197,198..989)
                     /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(347..366)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1160..1748
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1649..1668
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1902..1919
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2036..2057
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2072..2077,2078..2095,2096..2102)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2110..2126
                     /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     2115..2137
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2134..2150
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2134..2150
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2169..2189
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2171..2189
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     CDS             2444..5089
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae GAL4"
                     /product="GAL4 transcriptional activator"
                     /label=GAL4
                     /translation="MKLLSSIEQACDICRLKKLKCSKEKPKCAKCLKNNWECRYSPKTK
                     RSPLTRAHLTEVESRLERLEQLFLLIFPREDLDMILKMDSLQDIKALLTGLFVQDNVNK
                     DAVTDRLASVETDMPLTLRQHRISATSSSEESSNKGQRQLTVSIDSAAHHDNSTIPLDF
                     MPRDALHGFDWSEEDDMSDGLPFLKTDPNNNGFFGDGSLLCILRSIGFKPENYTNSNVN
                     RLPTMITDRYTLASRSTTSRLLQSYLNNFHPYCPIVHSPTLMMLYNNQIEIASKDQWQI
                     LFNCILAIGAWCIEGESTDIDVFYYQNAKSHLTSKVFESGSIILVTALHLLSRYTQWRQ
                     KTNTSYNFHSFSIRMAISLGLNRDLPSSFSDSSILEQRRRIWWSVYSWEIQLSLLYGRS
                     IQLSQNTISFPSSVDDVQRTTTGPTIYHGIIETARLLQVFTKIYELDKTVTAEKSPICA
                     KKCLMICNEIEEVSRQAPKFLQMDISTTALTNLLKEHPWLSFTRFELKWKQLSLIIYVL
                     RDFFTNFTQKKSQLEQDQNDHQSYEVKRCSIMLSDAAQRTVMSVSSYMDNHNVTPYFAW
                     NCSYYLFNAVLVPIKTLLSNSKSNAENNETAQLLQQINTVLMLLKKLATFKIQTCEKYI
                     QVLEEVCAPFLLSQCAIPLPHISYNNSNGSAIKNIVGSATIAQYPTLPEENVNNISVKY
                     VSPGSVGPSPVPLKSGASFSDLVKLLSNRPPSRNSPVTIPRSTPSHRSVTPFLGQQQQL
                     QSLVPLTPSALFGGANFNQSGNIADSSLSFTFTNSSNGPNLITTQTNSQALSQPIASSN
                     VHDNFMNNEITASKIDDGNNSKPLSPGWTDQTAYNAFGITTGMFNTTTMDDVYNYLFDD
                     EDTPPNPKKE"
     primer_bind     complement(5585..5601)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5585..5601)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    complement(5610..5643)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(5746..5765)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             5757..6566
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(5824..5843)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     protein_bind    complement(6641..6674)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(6741..6760)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6742..6760)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6770..6787)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6770..6786)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6779..6801)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      complement(6927..7383)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7065..7086)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7277..7296
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
       61 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
      121 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
      181 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
      241 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
      301 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
      361 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
      421 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
      481 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
      541 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
      601 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
      661 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
      721 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
      781 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
      841 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
      901 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
      961 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     1021 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     1081 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     1141 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     1201 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     1261 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc
     1321 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     1381 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     1441 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     1501 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     1561 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     1621 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     1681 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     1741 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     1801 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     1861 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
     1921 aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat
     1981 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg
     2041 tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt
     2101 tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg
     2161 ccaagcgcgc aattaaccct cactaaaggg aacaaaagct ggccactgtt taaacttggc
     2221 gcgcctttgc gatcgcagac ctaggaaagc tagctatacg cgttataccg gtatcctgca
     2281 ggtagcatgc aagagatctt ataggcctcg agttaacgtt cgaggtcgac tctagataaa
     2341 gctttagagc tcttcttaag aaggatcccg taagcaaata aacaagcgca gctgaacaag
     2401 ctaaacaatc tgcagcccaa gcttgaagca agcctcctga aagatgaagc tactgtcttc
     2461 tatcgaacaa gcatgcgata tttgccgact taaaaagctc aagtgctcca aagaaaaacc
     2521 gaagtgcgcc aagtgtctga agaacaactg ggagtgtcgc tactctccca aaaccaaaag
     2581 gtctccgctg actagggcac atctgacaga agtggaatca aggctagaaa gactggaaca
     2641 gctatttcta ctgatttttc ctcgagaaga ccttgacatg attttgaaaa tggattcttt
     2701 acaggatata aaagcattgt taacaggatt atttgtacaa gataatgtga ataaagatgc
     2761 cgtcacagat agattggctt cagtggagac tgatatgcct ctaacattga gacagcatag
     2821 aataagtgcg acatcatcat cggaagagag tagtaacaaa ggtcaaagac agttgactgt
     2881 atcgattgac tcggcagctc atcatgataa ctccacaatt ccgttggatt ttatgcccag
     2941 ggatgctctt catggatttg attggtctga agaggatgac atgtcggatg gcttgccctt
     3001 cctgaaaacg gaccccaaca ataatgggtt ctttggcgac ggttctctct tatgtattct
     3061 tcgatctatt ggctttaaac cggaaaatta cacgaactct aacgttaaca ggctcccgac
     3121 catgattacg gatagataca cgttggcttc tagatccaca acatcccgtt tacttcaaag
     3181 ttatctcaat aattttcacc cctactgccc tatcgtgcac tcaccgacgc taatgatgtt
     3241 gtataataac cagattgaaa tcgcgtcgaa ggatcaatgg caaatccttt ttaactgcat
     3301 attagccatt ggagcctggt gtatagaggg ggaatctact gatatagatg ttttttacta
     3361 tcaaaatgct aaatctcatt tgacgagcaa ggtcttcgag tcaggttcca taattttggt
     3421 gacagcccta catcttctgt cgcgatatac acagtggagg cagaaaacaa atactagcta
     3481 taattttcac agcttttcca taagaatggc catatcattg ggcttgaata gggacctccc
     3541 ctcgtccttc agtgatagca gcattctgga acaaagacgc cgaatttggt ggtctgtcta
     3601 ctcttgggag atccaattgt ccctgcttta tggtcgatcc atccagcttt ctcagaatac
     3661 aatctccttc ccttcttctg tcgacgatgt gcagcgtacc acaacaggtc ccaccatata
     3721 tcatggcatc attgaaacag caaggctctt acaagttttc acaaaaatct atgaactaga
     3781 caaaacagta actgcagaaa aaagtcctat atgtgcaaaa aaatgcttga tgatttgtaa
     3841 tgagattgag gaggtttcga gacaggcacc aaagttttta caaatggata tttccaccac
     3901 cgctctaacc aatttgttga aggaacaccc ttggctatcc tttacaagat tcgaactgaa
     3961 gtggaaacag ttgtctctta tcatttatgt attaagagat tttttcacta attttaccca
     4021 gaaaaagtca caactagaac aggatcaaaa tgatcatcaa agttatgaag ttaaacgatg
     4081 ctccatcatg ttaagcgatg cagcacaaag aactgttatg tctgtaagta gctatatgga
     4141 caatcataat gtcaccccat attttgcctg gaattgttct tattacttgt tcaatgcagt
     4201 cctagtaccc ataaagactc tactctcaaa ctcaaaatcg aatgctgaga ataacgagac
     4261 cgcacaatta ttacaacaaa ttaacactgt tctgatgcta ttaaaaaaac tggccacttt
     4321 taaaatccag acttgtgaaa aatacattca agtactggaa gaggtatgtg cgccgtttct
     4381 gttatcacag tgtgcaatcc cattaccgca tatcagttat aacaatagta atggtagcgc
     4441 cattaaaaat attgtcggtt ctgcaactat cgcccaatac cctactcttc cggaggaaaa
     4501 tgtcaacaat atcagtgtta aatatgtttc tcctggctca gtagggcctt cacctgtgcc
     4561 attgaaatca ggagcaagtt tcagtgatct agtcaagctg ttatctaacc gtccaccctc
     4621 tcgtaactct ccagtgacaa taccaagaag cacaccttcg catcgctcag tcacgccttt
     4681 tctagggcaa cagcaacagc tgcaatcatt agtgccactg accccgtctg ctttgtttgg
     4741 tggcgccaat tttaatcaaa gtgggaatat tgctgatagc tcattgtcct tcactttcac
     4801 taacagtagc aacggtccga acctcataac aactcaaaca aattctcaag cgctttcaca
     4861 accaattgcc tcctctaacg ttcatgataa cttcatgaat aatgaaatca cggctagtaa
     4921 aattgatgat ggtaataatt caaaaccact gtcacctggt tggacggacc aaactgcgta
     4981 taacgcgttt ggaatcacta cagggatgtt taataccact acaatggatg atgtatataa
     5041 ctatctattc gatgatgaag ataccccacc aaacccaaaa aaagagtaaa atgaatcgta
     5101 gatactgaaa aaccccgcaa gttcacttca actgtgcatc gtgcaccatc tcaatttctt
     5161 tcatttatac atcgttttgc cttcttttat gtaactatac tcctctaagt ttcaatcttg
     5221 gccatgtaac ctctgatcta tagaattttt taaatgacta gaattaatgc ccatcttttt
     5281 tttggaccta aattcttcat gaaaatatat tacgagggct tattcagaag cttatcgata
     5341 ccgtcgacta aagccaaata gaaaattatt cagttcctgg cttaagtttt taaaagtgat
     5401 attatttatt tggttgtaac caaccaaaag aatgtaaata actaatacat aattatgtta
     5461 gttttaagtt agcaacaaat tgattttagc tatattagct acttggttaa taaatagaat
     5521 atatttattt aaagataatt gcgtttttat tgtcagggag tgagtttgct taaaaactcg
     5581 tttagatcca ctagttctag agcggccgca taacttcgta tagcatacat tatacgaagt
     5641 tatggcagct ctggcccgtg tctcaaaatc tctgatgtta cattgcacaa gataaaaata
     5701 tatcatcatg aacaataaaa ctgtctgctt acataaacag taatacaagg ggtgttatga
     5761 gccatattca acgggaaacg tcgaggccgc gattaaattc caacatggat gctgatttat
     5821 atgggtataa atgggctcgc gataatgtcg ggcaatcagg tgcgacaatc tatcgcttgt
     5881 atgggaagcc cgatgcgcca gagttgtttc tgaaacatgg caaaggtagc gttgccaatg
     5941 atgttacaga tgagatggtc agactaaact ggctgacgga atttatgcct cttccgacca
     6001 tcaagcattt tatccgtact cctgatgatg catggttact caccactgcg atccccggaa
     6061 aaacagcatt ccaggtatta gaagaatatc ctgattcagg tgaaaatatt gttgatgcgc
     6121 tggcagtgtt cctgcgccgg ttgcattcga ttcctgtttg taattgtcct tttaacagcg
     6181 atcgcgtatt tcgtctcgct caggcgcaat cacgaatgaa taacggtttg gttgatgcga
     6241 gtgattttga tgacgagcgt aatggctggc ctgttgaaca agtctggaaa gaaatgcata
     6301 aacttttgcc attctcaccg gattcagtcg tcactcatgg tgatttctca cttgataacc
     6361 ttatttttga cgaggggaaa ttaataggtt gtattgatgt tggacgagtc ggaatcgcag
     6421 accgatacca ggatcttgcc atcctatgga actgcctcgg tgagttttct ccttcattac
     6481 agaaacggct ttttcaaaaa tatggtattg ataatcctga tatgaataaa ttgcagtttc
     6541 atttgatgct cgatgagttt ttctaatcag aattggttaa ttggttgtaa cactggcaga
     6601 gcattacgct gacttgacgg gacggcgcaa gctcatgacc ataacttcgt atagcataca
     6661 ttatacgaag ttatgcagtc atgcgtctga gaattccact taattaagaa ggccggccct
     6721 tcctcagcgt ggccaattcg ccctatagtg agtcgtatta cgcgcgctca ctggccgtcg
     6781 ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac
     6841 atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac
     6901 agttgcgcag cctgaatggc gaatggaaat tgtaagcgtt aatattttgt taaaattcgc
     6961 gttaaatttt tgttaaatca gctcattttt ttaaccaata ggccgaaatc ggcaaaatcc
     7021 cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga
     7081 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg
     7141 atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag
     7201 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga
     7261 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg
     7321 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg
     7381 cgtcag
//
