LOCUS       sequence-403220-        7946 bp    DNA     circular SYN 15-APR-2025
DEFINITION  GFP-Fray under actin promoter for cell culture.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7946)
  AUTHORS   Jenny Lab
  TITLE     Jenny Lab Wnk Plasmids 2023
  JOURNAL   unpublished
REFERENCE   2  (bases 1 to 7946)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "unpublished"
COMMENT     Sequence Label: pAGW-Fray
FEATURES             Location/Qualifiers
     source          1..7946
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          106..171
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             301..321
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    746..880
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(783..802)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     837..856
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             join(977..1045,1046..1837)
                     /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(1195..1214)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2008..2596
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2497..2516
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2750..2767
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        2918..5427
                     /label=Ac5 promoter
                     /note="Drosophila melanogaster actin 5C promoter"
     regulatory      5465..5474
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(5471..5473,5474..5476,5477..6187)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(5471..5473,5474..5476,5477..6187)
                     /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(5516..5537)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5777..5796)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6124..6145
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    6197..6221
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(join(7936..7946,1..14))
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
ORIGIN
        1 ttgtacaaag tggtgacgta agctagcagg atctttgtga aggaacctta cttctgtggt
       61 gtgacataat tggacaaact acctacagag atttaaagct ctaaggtaaa tataaaattt
      121 ttaagtgtat aatgtgttaa actactgatt ctaattgttt gtgtatttta gattccaacc
      181 tatggaactg atgaatggga gcagtggtgg aatgccttta atgaggaaaa cctgttttgc
      241 tcagaagaaa tgccatctag tgatgatgag gctactgctg actctcaaca ttctactcct
      301 ccaaaaaaga agagaaaggt agaagacccc aaggactttc cttcagaatt gctaagtttt
      361 ttgagtcatg ctgtgtttag taatagaact cttgcttgct ttgctattta caccacaaag
      421 gaaaaagctg cactgctata caagaaaatt atggaaaaat attctgtaac ctttataagt
      481 aggcataaca gttataatca taacatactg ttttttctta ctccacacag gcatagagtg
      541 tctgctatta ataactatgc tcaaaaattg tgtaccttta gctttttaat ttgtaaaggg
      601 gttaataagg aatatttgat gtatagtgcc ttgactagag atcataatca gccataccac
      661 atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca
      721 taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata
      781 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
      841 tttgtccaaa ctcatcaatg tatcttatca tgtctggatc ccgtttaaac tacgcgtaat
      901 tcaaacaggg ttctggcgtc gttctcgtac tgttttcccc aggccagtgc tttagcgtta
      961 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     1021 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     1081 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     1141 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     1201 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     1261 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     1321 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     1381 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     1441 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     1501 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
     1561 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     1621 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     1681 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     1741 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     1801 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     1861 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     1921 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     1981 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     2041 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     2101 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     2161 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     2221 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     2281 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     2341 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     2401 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     2461 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     2521 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     2581 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     2641 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     2701 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     2761 gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat
     2821 tcattaatgc aggcaactcg tgaaaggtag gcggatcagc ggccgcagca tgcaattcta
     2881 tattctaaaa acacaaatga tacttctaaa aaaaaatcat gaatggcatc aactctgaat
     2941 caaatctttg cagatgcacc tacttctcat ttccactgtc acatcatttt tccagatctc
     3001 gctgcctgtt atgtggccca caaaccaaga cacgttttat ggccattaaa gctggctgat
     3061 cgtcgccaaa caccaaatac ataatgaata tgtacacatt cgagaaagaa gcgatcaaag
     3121 aagcgtcttc gggcggagta ggagaatgcg gaggagaagg agaacgagct gatctagtat
     3181 ctctccacaa tccaatgcca actgaccaac tggccatatt cggagcaatt tgaagccaat
     3241 ttccatcgcc tggcgatcgc tccattcttg gctatatgtt tttcaccgtt acccggggcc
     3301 attttcaaag actcgtcggc aagataagat tgtgtcactc gctgtctctc ttcatttgtc
     3361 gaagaatgct gaggaatttc gcgatgacgt cggcgagtat tttgaagaat gagaataatt
     3421 tgtatttata cgaaaatcag ttagtggaat tttctacaaa aacatgttat ctatagataa
     3481 ttttgttgca aaatatgttg actatgacaa agattgtatg tatatacctt taatgtattc
     3541 tcattttctt atgtatttat aatggcaatg atgatactga tgatatttta agatgatgcc
     3601 agaccaaaag gcttgaattt ctgcgtcttt tgccgaacgc agtgcatgtg caattgttgt
     3661 tttttggaat attcaatttt cggactgtcc gctttgattt cagtttcttg gcttattcaa
     3721 aaagcaaagt aaagccaaaa aagcgagatg gcaataccaa atgcggcaaa acggtagtgg
     3781 aaggaaaggg gtgcggggca gcggaaggaa gggtggggcg gggcgtggcg gggtctgtgg
     3841 ctgggcgcga cgtcaccgac gttggagcca ctcctttgac catgtgtgcg tgtgtgtatt
     3901 attcgtgtct cgccactcgc cggttgtttt tttcttttta tgctgcgctc tctctagcgc
     3961 catctcgctt acgcatgctc aacgcaccgc atgttgccgt ttccttttat gcgtcatttt
     4021 ggctcgaaat aggcaattat ttaaacaaag attagtcaac gaaaacgcta aaataaataa
     4081 gtctacaata tggttactta ttgccatgtg tgtgcagcca acgatagcaa caaaagcaac
     4141 aacacaggtg gctttccctc tttcactttt tgtttgcaag ccgcgtgcga gcaagacggc
     4201 acgaccggca aacgcaatta cgctgacaaa gagcagacga agttttggcg aaaaacatca
     4261 aggcgcctga tacgaatgca tttgcaataa caattgcgat atttaatatt gtttatgaag
     4321 ctgtttgact tcaaaacaca caaaaaaaaa aataaaacaa attatttgaa agagaattag
     4381 gaatcggacg cttatcgtta gggtaacaac aagaaatgct tactgagtca cagcctctgg
     4441 aaaactgccg caagccagag agagagagaa aaagagggag agcagcttag accgcatgtg
     4501 cttgtgtgtg aggcgtctct ctcttcgtct ctgttgcgca aacgcataga ctgcactgaa
     4561 aaaatcgatt acctattttt tatgaatgaa tatttgcact attactattc aaaactatta
     4621 agatagcaat cacattcaat agccaaatac tataccacct gagcgatgca acgaaatgat
     4681 caatttgagc aaaaatgctg catatttagg acggcatcat tatagaaatg cttcttgctg
     4741 tgtacttttc tctcgtctgg cagctgtttc gccgttattg ttaaaaccgg cttaagttag
     4801 gtgtgttttc tacgactagt gaatgcccta ctagaagatg tgtgttgcac aaaatgtccc
     4861 tggaataacc aatttgaagt gcagatagca gtaaacgtaa gctaatatga atattattta
     4921 actgtaatgt tttaatatcg ctggacatta ctaataaacc cactataaac acatgtacat
     4981 atgtatgttt tggcatacaa tgagtagttg gggaaaaaat gtgtaaaagc accgtgacca
     5041 tcacagcata aagataacca gctgaagtat cgaatatgag taacccccaa attgaatcac
     5101 atgccgcaac tgataggacc catggaagta cactcttcat ggcgatatac aagacacaca
     5161 caagcacgaa cacccagttg cggaggaaat tctccgtaaa tgaaaaccca atcggcgaac
     5221 aattcatacc catatatggt aaaagttttg aacgcgactt gagagcggag agcattgcgg
     5281 ctgataaggt tttagcgcta agcgggcttt ataaaacggg ctgcgggacc agttttcata
     5341 tcactaccgt ttgagttctt gtgctgtgtg gatactcctc ccgacacaaa gccgctccat
     5401 cagccagcag tcgtctaatc cagagacccc ggatccagat atcgaggcct gtctagagaa
     5461 gcccgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
     5521 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga
     5581 tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc
     5641 ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga
     5701 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
     5761 caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg
     5821 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
     5881 cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa
     5941 gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt
     6001 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc
     6061 cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga
     6121 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct
     6181 gtacaagcac cggtatacaa gtttgtacaa aaaagcaggc tctttaaagg aaccaattca
     6241 gtcgactgga tccaccatga cctccatacc cgccaatctg tctagcaata atgttgctgg
     6301 tgcagcaacc ttaccaggag ctgcggcacc gccagagaaa tatacatggc cgaactccaa
     6361 ggacgattac gagctgaggg acgtcattgg tgtgggtgcc acagccgttg tccatggggc
     6421 ttactgtatt ccaaggaacg agaagtgcgc cattaagcgc atcaacctgg agaagtggaa
     6481 cacatcgatg gacgagctgc tcaaggagat tcaggccatg tcctcctgtt ttcacgagaa
     6541 cgtggtgacc tatcacacct catttgtggt tagggaagag ctctggttgg tgctaagact
     6601 tctagaaggc ggttctttgc tggacatcat caagcacaag atgcgaacgt ccaactgcaa
     6661 acagggtgtc ttcgacgaag ccaccattgc cacggtactg aaggaagtcc taaagggatt
     6721 ggaatatttc cattcgaacg ggcaaatcca tcgtgatatc aaggccggca atattttgat
     6781 tggtgacgat ggcaccattc aaatcgctga cttcggtgtg agtgcatggc tggccactgg
     6841 cagggatttg tccaggcaga aagtgcgcca cacctttgtg ggcacaccgt gttggatggc
     6901 tcccgaggtg atggagcagg atcatggcta tgactttaag gcggacattt ggtcctttgg
     6961 aataacagct atcgaaatgg ccacgggcac tgcaccatac cataaatacc cgcccatgaa
     7021 ggtgctaatg ctgaccctgc aaaatgatcc gcccaccttg gacacgggtg cggacgataa
     7081 agatcaatat aaggcgtatg gcaagacatt ccgcaagatg atcgtcgagt gcttgcaaaa
     7141 ggagccatcg aaacgtccca cggctagtga gcttttgaag cacgctttct tcaagaaggc
     7201 caaggatcgc aaatatctca cccagacgtt gctgcaatct gggcccagta tggagaccag
     7261 ggttcacaag gcggcgaaac gccaaccagg agcctcgggc cgtttgcatc gcacggtgac
     7321 aggcgaatgg gtctggtcta gcgaagagga ggacaatggt ggctctgcca ccggttccgg
     7381 aacaggagac agaaaacatc catcttcgga ctcggattcc gaggatagac ccatgaatcg
     7441 attggaaaga gccgactcat cggacagcga ccgggaagag ccttctcccg aaataaccca
     7501 cagtgtttcc tcagccacag tgacgcctgg agctccagct gctgctggtg ctggtgcagc
     7561 ccaagagcta acagctggca ttgctcaatt accactacct agcgaagcgg ccggcgaggc
     7621 gccaccagtc aatcttgtcc ttcgcatgcg caacttgcga cgtgaacttc acgacattcg
     7681 cttcgagttc gttgtgggca aggacactgc cgagggtatc gctacagaac tggtagacgc
     7741 cggactcgtt gatgcgctgg atacccaacc gatggcaacg catctggatc aactgatagc
     7801 tgccagtgca acgatgaaaa cgattacttt ccaattgagc tccggcgtgc agccgggcga
     7861 agtgcccgac gaacgatcac tggtgggata tgcgcagatc tccatcacgg actagtgtcg
     7921 actcgagata tctagaccca gctttc
//
