LOCUS       sequence-290390-        6298 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of HsNanos2-Y12E.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6298)
  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 6298)
  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-HsNanos2-Y12E_X
FEATURES             Location/Qualifiers
     source          1..6298
                     /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(1854..1872)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1855..1872)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1892..1909)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1898..2122
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2168..2596
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2255..2274)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2465..2486
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2605..2625)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2610..2939
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2790..2925
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2852..2871
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3006..3800
                     /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(3060..3079)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3670..3689
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3974..4095
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4011..4030)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4065..4084
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4144..4160)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4144..4160)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4157..4179)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4168..4184
                     /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(4192..4198,4199..4216,4217..4222))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4237..4258
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4375..4392)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4546..5131)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4626..4645)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5302..6093,6094..6162))
                     /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     5925..5944
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6163..6267)
                     /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 gcagctgccc ccattcgaca tgtggaagga cgagttcaac
     1441 ctgagccagg tcgtgtgggc cctgatcgcc agcagaggcc agagactgga aacccaggaa
     1501 atcgaggaac ccagccctgg ccctccactg ggacaggatc agggactggg agcccctggc
     1561 gctaatggcg gactgggcac cctgtgcaat ttctgcaagc acaacggcga gagccggcac
     1621 gtgtacagca gccaccagct gaaaaccccc gacggcgtgg tcgtgtgccc catcctgaga
     1681 cactacgtgt gccctgtgtg cggcgccact ggggatcagg cccataccct gaagtactgc
     1741 cccctgaatg gcggccagca gtccctgtac agaagaagcg gcagaaacag cgctgggcgg
     1801 agagtgaaga gataatctag agtcgacccg ggcggccgcc tagagggccc tattctatag
     1861 tgtcacctaa atgctagagc tcgctgatca gcctcgactg tgccttctag ttgccagcca
     1921 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
     1981 ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg
     2041 gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct
     2101 ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg
     2161 tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     2221 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     2281 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     2341 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     2401 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     2461 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     2521 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     2581 aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
     2641 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
     2701 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     2761 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     2821 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg
     2881 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
     2941 agctcccggg agcttgtata tccattttcg gatctgatca agagacagga tgaggatcgt
     3001 ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc
     3061 tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc
     3121 tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg
     3181 aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag
     3241 ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg
     3301 ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg
     3361 caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac
     3421 atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg
     3481 acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc
     3541 ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg
     3601 aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc
     3661 aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc
     3721 gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc
     3781 ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc
     3841 caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg
     3901 aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt
     3961 cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     4021 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     4081 catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc
     4141 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     4201 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     4261 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     4321 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     4381 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     4441 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     4501 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     4561 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     4621 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     4681 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     4741 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     4801 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     4861 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     4921 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     4981 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     5041 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtttttttg tttgcaagca
     5101 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     5161 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     5221 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     5281 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     5341 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     5401 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     5461 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     5521 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     5581 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     5641 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     5701 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     5761 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     5821 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     5881 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     5941 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     6001 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     6061 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     6121 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     6181 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     6241 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtc
//
