LOCUS       sequence-290406-        5108 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian Expression of HsNanos1-Y46E.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5108)
  AUTHORS   Bhandari D, Raisch T, Weichenrieder O, Jonas S, Izaurralde E
  TITLE     Structural basis for the Nanos-mediated recruitment of the CCR4-NOT 
            complex and translational repression.
  JOURNAL   Genes Dev. 2014 Apr 15;28(8):888-901. doi: 10.1101/gad.237289.113.
  PUBMED    24736845
REFERENCE   2  (bases 1 to 5108)
  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"; doi: 
            "10.1101/gad.237289"; journalName: "Genes Dev"; date: "2014-04-15- 
            15"; volume: "28"; issue: "8"; pages: "888-901"
COMMENT     Sequence Label: pT7-V5-SBP-C1-HsNanos1-Y46E_X
FEATURES             Location/Qualifiers
     source          1..5108
                     /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             668..709
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             731..844
                     /codon_start=1
                     /product="streptavidin-binding peptide"
                     /label=SBP
                     /note="selected from a peptide library; binds streptavidin 
                     with nanomolar affinity (Keefe et al., 2001)"
                     /translation="MDEKTTGWGGGHVVEGLAGELEQLRARLEHHPQGQREP"
     polyA_signal    1896..2017
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1933..1952)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1987..2006
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2024..2479)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2161..2182)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2373..2392
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2506..2610
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2612..2969
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2612..2803
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2635..2655)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2820..2955
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3004..3798
                     /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             3004..3798
                     /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(3058..3077)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3668..3687
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3986..4005)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4030..4077
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4406..4994
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4895..4914
                     /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 caccatgggt aagcctatcc ctaaccctct cctcggtctc gattctacgc ccggaagcgg
      721 tagcggtacg atggacgaga agaccaccgg ctgggggggc ggccacgtgg tggagggcct
      781 ggccggcgag ctggagcagc tgcgggccag gctggagcac caccctcagg gccagcggga
      841 gcccagatct cgagctatgg aagccttccc atgggccccc agatccccca gaagaggcag
      901 agcacctcct cccatggccc tggtgccaag cgccagatat gtgtctgccc ctggacctgc
      961 ccacccccag cctttcagca gctggaacga tgagctgggc ctggccaccc tgatcaccaa
     1021 ggccgtggat ggcgagccca gattcggatg tgccagaggc ggaaatggcg gcggaggcag
     1081 ccctcctagt agcagcagca gctcctgttg cagccctcac acaggcgctg gacctggcgc
     1141 tctgggacct gcactgggcc cacctgacta cgacgaggac gacgacgatg acagcgacga
     1201 gcctggcagc agaggcagat acctgggctc tgccctggaa ctgcgggctc tggaactgtg
     1261 tgccggacct gctgaagccg gcctgctgga agagagattc gccgagctga gccccttcgc
     1321 cggcagagct gcagcagtgc tgctgggatg tgctcctgct gccgcagccg ccgctacaac
     1381 aacatctgag gccacccctc gggaagagag ggctcctgct tgggccgctg aacctagact
     1441 gcatgctgcc tctggcgccg ctgctgcaag actgctgaaa cccgaactgc aagtgtgcgt
     1501 gttctgccgg aacaacaaag aagctatggc cctgtacacc acccacatcc tgaagggccc
     1561 cgacggcaga gtgctgtgtc ccgtgctgcg gagatacaca tgccctctgt gtggcgccag
     1621 cggcgacaat gcccacacca tcaagtactg ccccctgagc aaggtgcccc ctccaccagc
     1681 tagaccccct ccaagatctg ccagagatgg cccccctggc aagaagctga gataagaatt
     1741 ctgcagtcga cggtaccgcg ggcccgggat ccaccggatc tagataactg atcataatca
     1801 gccataccac atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga
     1861 acctgaaaca taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg
     1921 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     1981 ctagttgtgg tttgtccaaa ctcatcaatg tatcttaacg cgtaaattgt aagcgttaat
     2041 attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc
     2101 gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt
     2161 ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa
     2221 accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg
     2281 tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga
     2341 cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct
     2401 agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat
     2461 gcgccgctac agggcgcgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     2521 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     2581 atgcttcaat aatattgaaa aaggaagagt cctgaggcgg aaagaaccag ctgtggaatg
     2641 tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     2701 tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa
     2761 gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta actccgccca
     2821 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
     2881 ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag
     2941 gcttttttgg aggcctaggc ttttgcaaag atcgatcaag agacaggatg aggatcgttt
     3001 cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta
     3061 ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg
     3121 tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa
     3181 ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct
     3241 gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg
     3301 caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca
     3361 atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat
     3421 cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac
     3481 gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc
     3541 gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa
     3601 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag
     3661 gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc
     3721 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt
     3781 cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca
     3841 acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa
     3901 tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct
     3961 tcgcccaccc tagggggagg ctaactgaaa cacggaagga gacaataccg gaaggaaccc
     4021 gcgctatgac ggcaataaaa agacagaata aaacgcacgg tgttgggtcg tttgttcata
     4081 aacgcggggt tcggtcccag ggctggcact ctgtcgatac cccaccgaga ccccattggg
     4141 gccaatacgc ccgcgtttct tccttttccc caccccaccc cccaagttcg ggtgaaggcc
     4201 cagggctcgc agccaacgtc ggggcggcag gccctgccat agcctcaggt tactcatata
     4261 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     4321 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     4381 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     4441 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     4501 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag
     4561 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     4621 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     4681 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     4741 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     4801 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     4861 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     4921 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     4981 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     5041 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     5101 ccatgcat
//
