LOCUS       sequence-291395-        7883 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of HsGIGYF1-W294AF306AF312A.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7883)
  AUTHORS   Peter D, Ruscica V, Bawankar P, Weber R, Helms S, Valkov E, Igreja 
            C, Izaurralde E
  TITLE     Molecular basis for GIGYF-Me31B complex assembly in 4EHP-mediated 
            translational repression.
  JOURNAL   Genes Dev. 2019 Oct 1;33(19-20):1355-1360. doi: 
            10.1101/gad.329219.119. Epub 2019 Aug 22.
  PUBMED    31439631
REFERENCE   2  (bases 1 to 7883)
  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: "Genes 
            Dev."; date: "2019-10-1"; volume: "33(19-20):1355-1360. doi"; pages:
            " 10.1101/gad.329219.119. Epub 2019 Aug 22"
COMMENT     Sequence Label: pT7-EGFP-C1-HsGIGYF1-W294AF306AF312A_AI
FEATURES             Location/Qualifiers
     source          1..7883
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     613..632
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        613..631
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      659..668
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(665..667,668..670,671..1381)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(665..667,668..670,671..1381)
                     /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(710..731)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(971..990)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1318..1339
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    4671..4792
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4708..4727)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4762..4781
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4799..5254)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4936..4957)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5148..5167
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5281..5385
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5387..5744
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5387..5578
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5410..5430)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5595..5730
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5779..6573
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             5779..6573
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5833..5852)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6443..6462
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6761..6780)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6805..6852
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      7181..7769
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7670..7689
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca ctgcttactg
      601 gcttatcgaa attaatacga ctcactatag ggagacccaa gctctagcgc taccggtcgc
      661 caccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
      721 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac
      781 ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc
      841 caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat
      901 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat
      961 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac
     1021 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg
     1081 gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa
     1141 gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct
     1201 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa
     1261 ccactacctg agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat
     1321 ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa
     1381 gtccggactc agatctcgag ctatggcagc agagacactc aactttgggc ctgagtggct
     1441 cagggccctg tccgggggcg gcagcgtggc ctccccaccc ccgtcccctg ccatgcccaa
     1501 atacaagctg gctgactacc gttatgggcg agaggaaatg ctggctctct acgtcaagga
     1561 gaacaaggtc ccggaagagc tgcaggacaa ggagttcgcc gcggtgctgc aggacgagcc
     1621 actgcagccc ctggctctgg agccgctgac tgaggaggaa cagagaaact tctccctgtc
     1681 agtgaacagc gtggctgtgc tgaggctgat ggggaaaggg gctggccccc ccctggctgg
     1741 cacctcccga ggcaggggca gcacgcggag ccgaggccgc ggccgtggtg acagctgctt
     1801 ttaccaaaga agcatcgaag aaggcgatgg ggcctttgga cgaagccccc gggaaatcca
     1861 gcgcagccag agctgggatg acagaggcga gaggcggttt gagaagtcag caaggcggga
     1921 tggagcacga tgtggctttg aggagggagg ggctggccca aggaaggagc acgcccgctc
     1981 agacagcgag aactggcgct ccctacggga ggaacaggag gaggaggagg agggcagctg
     2041 gaggctcgga gcagggcccc ggcgagacgg cgaccgctgg cgctccgcca gccctgatgg
     2101 tggtccccgc tctgctggct ggcgggaaca tggggaacgg cggcgcaagt ttgaatttga
     2161 tttgcgaggg gatcgaggag ggtgtggtga agaggagggg cggggagggg gaggcagctc
     2221 tcacctgcgg cggtgccgag cgcctgaagg ctttgaggag gacaaggatg ggctcccaga
     2281 ggcgtgcctg gacgatgagg atgaagaaat gggcaccgcc gatgcctctg gggccgcctt
     2341 gcctctcaag aagggcccca aggagcccat tcctgaggag caggagctgg acttccaagg
     2401 gttggaggag gaggaggaac cttccgaagg gctagaggag gaagggcctg aggcaggtgg
     2461 gaaagagctg accccactgc ctcctcagga ggagaagtcc agctccccat ccccactgcc
     2521 caccctgggc ccactctggg ggacaaacgg ggatggggac gaaactgcag agaaagagcc
     2581 cccagcggcc gaagatgata ttcgggggat ccagctgagt cccggggtgg gctcctctgc
     2641 tggcccaccc ggagatctgg aggatgatga aggcttgaag cacctgcagc aggaggcgga
     2701 gaagctggtg gcctccctgc aggacagctc cttggaggag gagcagttca cggctgccat
     2761 gcagacccag ggcctgcgcc actctgcagc cgccactgcc ctcccgctca gccatggggc
     2821 tgcccggaag tggttctaca aggacccaca gggcgagatc caaggcccct tcacgacaca
     2881 ggagatggca gagtggttcc aggccggcta cttttccatg tcactgctgg tgaagcgggg
     2941 ctgcgatgag ggcttccagc cgctgggcga ggtgatcaag atgtggggcc gcgtgccctt
     3001 tgccccaggg ccctcacctc ccccactgct gggaaacatg gaccaggagc ggctgaagaa
     3061 gcaacaggag ctggccgcgg cggccttgta ccagcagctg cagcaccagc agtttctcca
     3121 gctggtcagc agccgccagc tcccacagtg cgcgctccga gaaaaggcag ctctggggga
     3181 cctgacaccg ccaccaccgc cgccgccaca gcagcagcag cagcagctca cggcattcct
     3241 gcagcagctc caggcgctca aaccccccag aggcggggac cagaacctgc tcccgacgat
     3301 gagccggtcc ttgtcggtgc cagattcggg ccgcctctgg gacgtacata cctcagcctc
     3361 atcacagtca ggtggtgagg ccagtctttg ggacatacca attaactctt cgactcaggg
     3421 tccaattcta gaacaactcc agctgcaaca taaattccag gagcgcagag aagtggagct
     3481 cagggcgaag cgggaggaag aggaacgcaa gcgtcgagag gagaagcgcc gccagcagca
     3541 gcaggaggag cagaagcggc ggcaggagga ggaagagctg tttcggcgca agcacgtgcg
     3601 gcagcaggag ctattgctga agttgctaca gcagcagcag gcggtccctg tgccccccgc
     3661 acccagctcc ccgcccccac tctgggctgg cctggccaag caggggctgt ccatgaagac
     3721 gctcctggag ttgcagctgg agggcgagcg gcagctgcac aaacagcccc cacctcggga
     3781 gccagctcgg gcccaggccc ccaaccaccg agtgcagctt gggggcctgg gcactgcccc
     3841 cctgaaccag tgggtgtctg aggctgggcc actgtggggc gggccagaca agagtggggg
     3901 cggcagcagc ggcctggggc tctgggagga cacccccaag agcggcggga gcctggtccg
     3961 tggcctcggc ctgaagaaca gccggagcag cccatctctc agtgactcat acagccacct
     4021 atcgggtcgg cccattcgca aaaagacgga ggaagaagag aagctgctga agctgctgca
     4081 gggcattccc aggccccagg acggcttcac ccagtggtgc gagcagatgc tgcacacgct
     4141 gagcgccacg ggcagcctgg acgtgcccat ggctgtagcg atcctcaagg aggtggaatc
     4201 cccctatgat gtccacgatt atatccgttc ctgcctgggg gacacgctgg aagccaaaga
     4261 atttgccaaa caattcctgg agcggagggc caagcagaaa gccagccagc agcggcagca
     4321 gcagcaggag gcatggctga gcagcgcctc gctgcagacg gccttccagg ccaaccacag
     4381 caccaaactc ggccccgggg agggcagcaa ggccaagagg cgggcactga tgctgcactc
     4441 agaccccagc atcctggggt actccctgca cggatcttct ggtgagatcg agagcgtgga
     4501 tgactactga gaattctgca gtcgacggta ccgcgggccc gggatccacc ggatctagat
     4561 aactgatcat aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc
     4621 cacacctccc cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta
     4681 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     4741 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct taacgcgtaa
     4801 attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt
     4861 tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata
     4921 gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac
     4981 gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa
     5041 tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc
     5101 cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg
     5161 aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca
     5221 cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg
     5281 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     5341 caataaccct gataaatgct tcaataatat tgaaaaagga agagtcctga ggcggaaaga
     5401 accagctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     5461 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     5521 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     5581 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     5641 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc
     5701 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaagatcga tcaagagaca
     5761 ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct
     5821 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
     5881 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
     5941 ggtgccctga atgaactgca agacgaggca gcgcggctat cgtggctggc cacgacgggc
     6001 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
     6061 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
     6121 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac
     6181 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
     6241 caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc
     6301 aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg
     6361 aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg
     6421 gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc
     6481 gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc
     6541 gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg
     6601 accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa
     6661 ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc
     6721 tcatgctgga gttcttcgcc caccctaggg ggaggctaac tgaaacacgg aaggagacaa
     6781 taccggaagg aacccgcgct atgacggcaa taaaaagaca gaataaaacg cacggtgttg
     6841 ggtcgtttgt tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac
     6901 cgagacccca ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa
     6961 gttcgggtga aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcct
     7021 caggttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     7081 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     7141 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     7201 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     7261 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     7321 atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     7381 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     7441 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     7501 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     7561 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     7621 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     7681 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     7741 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     7801 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     7861 ataaccgtat taccgccatg cat
//
