LOCUS       sequence-310839-        4390 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Donor for precise CRISPR directed genomic integration of QFGal4 
            using the GeneWeld method.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4390)
  AUTHORS   Burgess J, Burrows JT, Sadhak R, Chiang S, Weiss A, D'Amata C, 
            Molinaro AM, Zhu S, Long M, Hu C, Krause HM, Pearson BJ
  TITLE     An optimized QF-binary expression system for use in zebrafish.
  JOURNAL   Dev Biol. 2020 Jul 19. pii: S0012-1606(20)30202-5. doi: 
            10.1016/j.ydbio.2020.07.007.
  PUBMED    32697972
REFERENCE   2  (bases 1 to 4390)
  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: "Dev Biol. 
            2020 Jul 19. pii: S0012-1606(20)30202-5. doi: 
            10.1016/j.ydbio.2020.07.007."
COMMENT     Sequence Label: pGTag-P2A-QFGal4-SV40pA
FEATURES             Location/Qualifiers
     source          1..4390
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(157..184)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(276..362)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     426..442
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    525..546
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(561..566,567..584,585..591)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    599..615
                     /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     604..626
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     623..639
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             635..1024
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL
                     QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWTCHRIRSHELLS
                     VKASRRRGRKPRSRDGTQEEEEDAA"
     primer_bind     658..678
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        660..678
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        complement(726..744)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(754..770)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             930..986
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             1647..1991
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae GAL4 (truncated)"
                     /product="activation domain of the GAL4 transcriptional 
                     activator"
                     /label=GAL4 activation domain
                     /translation="ANFNQSGNIADSSLSFTFTNSSNGPNLITTQTNSQALSQPIASSN
                     VHDNFMNNEITASKIDDGNNSKPLSPGWTDQTAYNAFGITTGMFNTTTMDDVYNYLFDD
                     EDTPPNPKKE"
     polyA_signal    2030..2164
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2054..2073)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2108..2127
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     protein_bind    complement(2227..2322)
                     /gene="mutant version of attL"
                     /label=attL3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        complement(2521..2539)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2544..2560)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2662..2681)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2673..3482
                     /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(2740..2759)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3629..4217
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4118..4137
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4371..4388
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcggatgtg cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg
       61 cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatac
      121 gcgtaccgct agccaggaag agtttgtaga aacgcaaaaa ggccatccgt caggatggcc
      181 ttctgcttag tttgatgcct ggcagtttat ggcgggcgtc ctgcccgcca ccctccgggc
      241 cgttgcttca caacgttcaa atccgctccc ggcggatttg tcctactcag gagagcgttc
      301 accgacaaac aacagataaa acgaaaggcc cagtcttccg actgagcctt tcgttttatt
      361 tgatgcctgg cagttcccta ctctcgcgtt aacgctagca tggatgtttt cccagtcacg
      421 acgttgtaaa acgacggcca gtcttaagct cgggccctgc agctctagag ctcgaattcg
      481 ggaggcgttc gggccacagc ggacacgcag gtgcgcaacg caattaatgt gagttagctc
      541 actcattagg caccccaggc tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt
      601 gtgagcggat aacaatttca cacaggaaac agctatgacc atgattacgc caagcgcgca
      661 attaaccctc actaaaggga acaaaagctg gaggacctcg agggggggcc cggtacccaa
      721 ttcgccctat agtgagtcgt attacgcgcg ctcactggcc gtcgttttac aacgtcgtga
      781 ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag
      841 ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa
      901 tggcgaatgg acctgccaca ggatccggag ccacgaactt ctctctgtta aagcaagcag
      961 gagacgtgga agaaaacccc ggtcccggga tggcacccaa gaagaagagg aagatgccgc
     1021 ctaaacgcaa gacactcaat gccgctgccg aagccaatgc ccacgctgat ggccatgctg
     1081 atggcaatgc tgatggtcat gtcgctaaca ctgcagcaag cagcaacaac gcccgttttg
     1141 cggacttgac caacattgac acacccggcc tcggccctac cacgacgacg ttacttgtcg
     1201 agcccgctcg ttcgaaacgc cagagagtct cgagggcctg tgatcagtgt cgagctgcac
     1261 gtgaaaagtg tgatggaatc cagccggctt gcttcccctg tgtgtcgcag ggccggtcgt
     1321 gtacctacca ggccagtccc aagaagcgag gagtccagac gggctacatc cgcactctcg
     1381 aactggctct ggcttggatg ttcgagaacg ttgcccgcag cgaggacgcc ctccacaatc
     1441 ttttggtccg tgatgctggc cagggcagcg ctctcctggt cggcaaagac tcgcctgctg
     1501 cagaacgcct gcatgcaaga tgggcgacga gtcgagtcaa caaaagcatc acccgtcttc
     1561 tctcaggtgc tagcactagt agcgccggtc aacagcaaca gctgcaatca ttagtgccac
     1621 tgaccccgtc tgctttgttt ggtggcgcca attttaatca aagtgggaat attgctgata
     1681 gctcattgtc cttcactttc actaacagta gcaacggtcc gaacctcata acaactcaaa
     1741 caaattctca agcgctttca caaccaattg cctcctctaa cgttcatgat aacttcatga
     1801 ataatgaaat cacggctagt aaaattgatg atggtaataa ttcaaaacca ctgtcacctg
     1861 gttggacgga ccaaactgcg tataacgcgt ttggaatcac tacagggatg tttaatacca
     1921 ctacaatgga tgatgtatat aactatctat tcgatgatga agatacccca ccaaacccaa
     1981 aaaaagagta aaggcgcgcc tctagaacta tagtgagtcg tattacgtag atccagacat
     2041 gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt
     2101 tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca
     2161 agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggagg tgtgggaggt
     2221 tttttccaac tttattatac aaagttggca ttataaaaaa gcattgctta tcaatttgtt
     2281 gcaacgaaca ggtcactatc agtcaaaata aaatcattat ttggagctcc atggcgttgt
     2341 ctagcaagga agtgaagagc tatccggata tatatgtatg ttatcataac tgtactggct
     2401 atgtcaggtg gtaatgactg taacgttacg ttactcgttg taggcacgac attgaatatg
     2461 ctcttctccg ctgtggcccg aacgcctccc gctagcgatg ggatctcggc cgcgatatcc
     2521 cctatagtga gtcgtattac atggtcatag ctgtttcctg gcagctctgg cccgtgtctc
     2581 aaaatctctg atgttacatt gcacaagata aaaatatatc atcatgaaca ataaaactgt
     2641 ctgcttacat aaacagtaat acaaggggtg ttatgagcca tattcaacgg gaaacgtcga
     2701 ggccgcgatt aaattccaac atggatgctg atttatatgg gtataaatgg gctcgcgata
     2761 atgtcgggca atcaggtgcg acaatctatc gcttgtatgg gaagcccgat gcgccagagt
     2821 tgtttctgaa acatggcaaa ggtagcgttg ccaatgatgt tacagatgag atggtcagac
     2881 taaactggct gacggaattt atgcctcttc cgaccatcaa gcattttatc cgtactcctg
     2941 atgatgcatg gttactcacc actgcgatcc ccggaaaaac agcattccag gtattagaag
     3001 aatatcctga ttcaggtgaa aatattgttg atgcgctggc agtgttcctg cgccggttgc
     3061 attcgattcc tgtttgtaat tgtcctttta acagcgatcg cgtatttcgt ctcgctcagg
     3121 cgcaatcacg aatgaataac ggtttggttg atgcgagtga ttttgatgac gagcgtaatg
     3181 gctggcctgt tgaacaagtc tggaaagaaa tgcataaact tttgccattc tcaccggatt
     3241 cagtcgtcac tcatggtgat ttctcacttg ataaccttat ttttgacgag gggaaattaa
     3301 taggttgtat tgatgttgga cgagtcggaa tcgcagaccg ataccaggat cttgccatcc
     3361 tatggaactg cctcggtgag ttttctcctt cattacagaa acggcttttt caaaaatatg
     3421 gtattgataa tcctgatatg aataaattgc agtttcattt gatgctcgat gagtttttct
     3481 aatcagaatt ggttaattgg ttgtaacact ggcagagcat tacgctgact tgacgggacg
     3541 gcgcaagctc atgaccaaaa tcccttaacg tgagttacgc gtcgttccac tgagcgtcag
     3601 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     3661 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     3721 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc
     3781 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     3841 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     3901 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     3961 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     4021 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     4081 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     4141 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     4201 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     4261 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     4321 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag
     4381 tgagcgagga
//
