LOCUS       sequence-310829-        3866 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Middle entry vector containing QUAS5x element upstream of GFP-NLS..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3866)
  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 3866)
  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: pME-QUAS5x:GFPNLS-SV40pA
FEATURES             Location/Qualifiers
     source          1..3866
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     93..110
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(269..296)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(388..474)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     523..545
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     537..554
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     538..554
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    570..669
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     regulatory      927..936
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(933..935,936..938,939..1649)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(933..935,936..938,939..1649)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(978..999)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1239..1258)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1586..1607
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1832..1851)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1886..1905
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1949..1965)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1950..1966)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     protein_bind    complement(1992..2091)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        complement(2109..2127)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2132..2148)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2132..2148)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2250..2269)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2261..3070
                     /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(2328..2347)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3217..3805
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3706..3725
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
       61 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
      121 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
      181 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat acgcgtaccg
      241 ctagccagga agagtttgta gaaacgcaaa aaggccatcc gtcaggatgg ccttctgctt
      301 agtttgatgc ctggcagttt atggcgggcg tcctgcccgc caccctccgg gccgttgctt
      361 cacaacgttc aaatccgctc ccggcggatt tgtcctactc aggagagcgt tcaccgacaa
      421 acaacagata aaacgaaagg cccagtcttc cgactgagcc tttcgtttta tttgatgcct
      481 ggcagttccc tactctcgcg ttaacgctag catggatgtt ttcccagtca cgacgttgta
      541 aaacgacggc cagtcttaag ctcgggcccc aaataatgat tttattttga ctgatagtga
      601 cctgttcgtt gcaacaaatt gatgagcaat gcttttttat aatgccaact ttgtacaaaa
      661 aagcaggctc gaattggagc tccaccgcgg tggcggccgc aaggataaac aattatcctc
      721 aaggataaac aattatcctc aaggataaac aattatcctc aaggataaac aattatcctc
      781 aaggataaac aattatcctc gcaagggtcg actctagagg gtatataatg gatcccatcg
      841 cgtctcagcc tcactttgag ctcctccaca cgaattccct cgacctcgaa gacgcgtctg
      901 tttgggatcc accggtcact agtgccgcca ccatggtgag caagggcgag gagctgttca
      961 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
     1021 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca
     1081 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc
     1141 agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
     1201 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc
     1261 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
     1321 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca
     1381 acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc
     1441 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     1501 gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca
     1561 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     1621 tcactctcgg catggacgag ctgtacaaga gcaggcacag aaggcatcgc cagcgctcta
     1681 ggagccgcaa tcgcagccga agtcgcagca gtgaacgaaa acgccgtcaa cggagccgaa
     1741 gtcgcagcag tgaacgaaga cgctacttgt acaagtaatc tagaactata gtgagtcgta
     1801 ttacgtagat ccagacatga taagatacat tgatgagttt ggacaaacca caactagaat
     1861 gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat
     1921 tataagctgc aataaacaag aagcttatcg ataccgtcga cctcgagggg gggcccggta
     1981 cccagctttt gacccagctt tcttgtacaa agttggcatt ataagaaagc attgcttatc
     2041 aatttgttgc aacgaacagg tcactatcag tcaaaataaa atcattattt gccatccagc
     2101 tgatatcccc tatagtgagt cgtattacat ggtcatagct gtttcctggc agctctggcc
     2161 cgtgtctcaa aatctctgat gttacattgc acaagataaa aatatatcat catgaacaat
     2221 aaaactgtct gcttacataa acagtaatac aaggggtgtt atgagccata ttcaacggga
     2281 aacgtcgagg ccgcgattaa attccaacat ggatgctgat ttatatgggt ataaatgggc
     2341 tcgcgataat gtcgggcaat caggtgcgac aatctatcgc ttgtatggga agcccgatgc
     2401 gccagagttg tttctgaaac atggcaaagg tagcgttgcc aatgatgtta cagatgagat
     2461 ggtcagacta aactggctga cggaatttat gcctcttccg accatcaagc attttatccg
     2521 tactcctgat gatgcatggt tactcaccac tgcgatcccc ggaaaaacag cattccaggt
     2581 attagaagaa tatcctgatt caggtgaaaa tattgttgat gcgctggcag tgttcctgcg
     2641 ccggttgcat tcgattcctg tttgtaattg tccttttaac agcgatcgcg tatttcgtct
     2701 cgctcaggcg caatcacgaa tgaataacgg tttggttgat gcgagtgatt ttgatgacga
     2761 gcgtaatggc tggcctgttg aacaagtctg gaaagaaatg cataaacttt tgccattctc
     2821 accggattca gtcgtcactc atggtgattt ctcacttgat aaccttattt ttgacgaggg
     2881 gaaattaata ggttgtattg atgttggacg agtcggaatc gcagaccgat accaggatct
     2941 tgccatccta tggaactgcc tcggtgagtt ttctccttca ttacagaaac ggctttttca
     3001 aaaatatggt attgataatc ctgatatgaa taaattgcag tttcatttga tgctcgatga
     3061 gtttttctaa tcagaattgg ttaattggtt gtaacactgg cagagcatta cgctgacttg
     3121 acgggacggc gcaagctcat gaccaaaatc ccttaacgtg agttacgcgt cgttccactg
     3181 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     3241 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     3301 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     3361 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     3421 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     3481 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     3541 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     3601 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     3661 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     3721 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     3781 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     3841 cttttgctgg ccttttgctc acatgt
//
