LOCUS       sequence-290412-        4751 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of HsNanos3-W7E.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4751)
  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 4751)
  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"; 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-HsNanos3-W7E_X
FEATURES             Location/Qualifiers
     source          1..4751
                     /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    1539..1660
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1576..1595)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1630..1649
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1667..2122)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1804..1825)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2016..2035
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2149..2253
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2255..2612
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2255..2446
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2278..2298)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2463..2598
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2647..3441
                     /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             2647..3441
                     /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(2701..2720)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3311..3330
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3629..3648)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3673..3720
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4049..4637
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4538..4557
                     /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 gcaccttcga cctggagacc gactatctgg gactggccca
      901 tctcgtgcgg gccctgtctg gaaaagaggg ccccgagaca agactgagcc cccagcctga
      961 gccagagccc atgctggaac ccgaccagaa gcggagcctg gaaagctctc ctgcccccga
     1021 gagactgtgc agcttctgca agcacaacgg cgagagccgg gccatctacc agagccacgt
     1081 gctgaaggat gaggccggca gagtgctgtg ccccatcctg agagactacg tgtgccctca
     1141 gtgcggcgcc accagagaga gagcccacac cagaagattc tgccccctga ccggccaggg
     1201 ctacaccagc gtgtacagcc acaccaccag aaacagcgcc ggcaagaaac tcgtgcggcc
     1261 cgacaaggcc aagacccagg ataccggaca cagaagaggc ggcggaggcg gagctggctt
     1321 tagaggcgcc ggaaagtctg agcccagccc cagctgtagc cccagcatga gcacctaaga
     1381 attctgcagt cgacggtacc gcgggcccgg gatccaccgg atctagataa ctgatcataa
     1441 tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc
     1501 tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata
     1561 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     1621 attctagttg tggtttgtcc aaactcatca atgtatctta acgcgtaaat tgtaagcgtt
     1681 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
     1741 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt
     1801 gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga
     1861 aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg
     1921 gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct
     1981 tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc
     2041 gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt
     2101 aatgcgccgc tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg cggaacccct
     2161 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
     2221 taaatgcttc aataatattg aaaaaggaag agtcctgagg cggaaagaac cagctgtgga
     2281 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     2341 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     2401 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     2461 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     2521 tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag
     2581 gaggcttttt tggaggccta ggcttttgca aagatcgatc aagagacagg atgaggatcg
     2641 tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg
     2701 ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg
     2761 ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat
     2821 gaactgcaag acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca
     2881 gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg
     2941 gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat
     3001 gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa
     3061 catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg
     3121 gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgagcatg
     3181 cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg
     3241 gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat
     3301 caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac
     3361 cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc
     3421 cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc
     3481 ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg
     3541 gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt
     3601 tcttcgccca ccctaggggg aggctaactg aaacacggaa ggagacaata ccggaaggaa
     3661 cccgcgctat gacggcaata aaaagacaga ataaaacgca cggtgttggg tcgtttgttc
     3721 ataaacgcgg ggttcggtcc cagggctggc actctgtcga taccccaccg agaccccatt
     3781 ggggccaata cgcccgcgtt tcttcctttt ccccacccca ccccccaagt tcgggtgaag
     3841 gcccagggct cgcagccaac gtcggggcgg caggccctgc catagcctca ggttactcat
     3901 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     3961 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     4021 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     4081 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     4141 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc
     4201 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     4261 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     4321 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     4381 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     4441 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     4501 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     4561 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     4621 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     4681 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     4741 ccgccatgca t
//
