LOCUS       sequence-290383-        6744 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of HsNanos1-W43E.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6744)
  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 6744)
  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: pcDNA3.1-MS2-HA-HsNanos1-W43E_X
FEATURES             Location/Qualifiers
     source          1..6744
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      895..904
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1333..1359
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     promoter        complement(2300..2318)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2301..2318)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2338..2355)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2344..2568
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2614..3042
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2701..2720)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2911..2932
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3051..3071)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3056..3385
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3236..3371
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3298..3317
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3452..4246
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3506..3525)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4116..4135
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4420..4541
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4457..4476)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4511..4530
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4590..4606)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4590..4606)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4603..4625)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4614..4630
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4638..4644,4645..4662,4663..4668))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4683..4704
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4821..4838)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4992..5577)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5072..5091)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5748..6539,6540..6608))
                     /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     6371..6390
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6609..6713)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttgccacc
      901 atggcttcta actttactca gttcgttctc gtcgacaatg gcggaactgg cgacgtgact
      961 gtcgccccaa gcaacttcgc taacggggtc gctgaatgga tcagctctaa ctcgcgatca
     1021 caggcttaca aagtaacctg tagcgttcgt cagagctctg cgcagaatcg caaatacacc
     1081 atcaaagtcg aggtgcctaa agtggcaacc cagactgttg gtggtgaaga gcttcctgta
     1141 gccggatgga gatcttactt aaatatggaa ctaaccattc caattttcgc cacgaattcc
     1201 gactgcgagc ttattgttaa ggcaatgcaa ggtctcctaa aagatggaaa cccgattccc
     1261 tcggccatcg cggccaactc cggcatctac ggaggtggag gtggatctgg tggatctcct
     1321 ccacatatgg cttacccata cgatgttcca gattacgctc ctccaaagct tggatccgaa
     1381 ttcccgggtc gactcgagat ggaagccttc ccatgggccc ccagatcccc cagaagaggc
     1441 agagcacctc ctcccatggc cctggtgcca agcgccagat atgtgtctgc ccctggacct
     1501 gcccaccccc agcctttcag cagcgagaac gattacctgg gcctggccac cctgatcacc
     1561 aaggccgtgg atggcgagcc cagattcgga tgtgccagag gcggaaatgg cggcggaggc
     1621 agccctccta gtagcagcag cagctcctgt tgcagccctc acacaggcgc tggacctggc
     1681 gctctgggac ctgcactggg cccacctgac tacgacgagg acgacgacga tgacagcgac
     1741 gagcctggca gcagaggcag atacctgggc tctgccctgg aactgcgggc tctggaactg
     1801 tgtgccggac ctgctgaagc cggcctgctg gaagagagat tcgccgagct gagccccttc
     1861 gccggcagag ctgcagcagt gctgctggga tgtgctcctg ctgccgcagc cgccgctaca
     1921 acaacatctg aggccacccc tcgggaagag agggctcctg cttgggccgc tgaacctaga
     1981 ctgcatgctg cctctggcgc cgctgctgca agactgctga aacccgaact gcaagtgtgc
     2041 gtgttctgcc ggaacaacaa agaagctatg gccctgtaca ccacccacat cctgaagggc
     2101 cccgacggca gagtgctgtg tcccgtgctg cggagataca catgccctct gtgtggcgcc
     2161 agcggcgaca atgcccacac catcaagtac tgccccctga gcaaggtgcc ccctccacca
     2221 gctagacccc ctccaagatc tgccagagat ggcccccctg gcaagaagct gagataagcg
     2281 gccgcctaga gggccctatt ctatagtgtc acctaaatgc tagagctcgc tgatcagcct
     2341 cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga
     2401 ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt
     2461 gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg
     2521 attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg
     2581 aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg
     2641 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     2701 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     2761 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     2821 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     2881 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     2941 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     3001 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg
     3061 tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     3121 tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     3181 gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc
     3241 gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat
     3301 ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt
     3361 ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc
     3421 tgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg
     3481 ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg
     3541 ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa
     3601 gaccgacctg tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct
     3661 ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga
     3721 ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc
     3781 cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac
     3841 ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc
     3901 cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact
     3961 gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga
     4021 tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg
     4081 ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga
     4141 agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga
     4201 ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg
     4261 ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc
     4321 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     4381 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat
     4441 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     4501 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg
     4561 acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     4621 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     4681 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     4741 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     4801 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     4861 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     4921 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     4981 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     5041 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     5101 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     5161 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     5221 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     5281 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     5341 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     5401 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     5461 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     5521 ggtagcggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     5581 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     5641 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     5701 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     5761 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     5821 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     5881 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     5941 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     6001 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     6061 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     6121 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     6181 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     6241 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     6301 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     6361 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     6421 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     6481 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     6541 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     6601 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     6661 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     6721 ccccgaaaag tgccacctga cgtc
//
