LOCUS       sequence-290275-        5128 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial Expression of HsNanos2_6-55.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5128)
  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 5128)
  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: pnYC-NpG-HsNanos2_6-22-Strep_W
FEATURES             Location/Qualifiers
     source          1..5128
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     2294..2313
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     2402..2421
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2402..2420
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2421..2445
                     /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)."
     RBS             2460..2482
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2489..3142
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MAPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     CDS             3164..3187
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             3260..3283
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     primer_bind     complement(3360..3378)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3374..3421
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(3596..3601,3602..3618,3619..3624))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     rep_origin      3736..4281
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     4182..4201
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4398..4415
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4807..4909
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             join(4910..5128,1..441)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFGMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(4976..4995)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
ORIGIN
        1 ggtatggcaa tgaaagacgg tgagctggtg atatgggata gtgttcaccc ttgttacacc
       61 gttttccatg agcaaactga aacgttttca tcgctctgga gtgaatacca cgacgatttc
      121 cggcagtttc tacacatata ttcgcaagat gtggcgtgtt acggtgaaaa cctggcctat
      181 ttccctaaag ggtttattga gaatatgttt ttcgtctcag ccaatccctg ggtgagtttc
      241 accagttttg atttaaacgt ggccaatatg gacaacttct tcgcccccgt tttcaccatg
      301 ggcaaatatt atacgcaagg cgacaaggtg ctgatgccgc tggcgattca ggttcatcat
      361 gccgtctgtg atggcttcca tgtcggcaga atgcttaatg aattacaaca gtactgcgat
      421 gagtggcagg gcggggcgta atttttttaa ggcagttatt ggtgccctta aacgcctggt
      481 gctacgcctg aataagtgat aataagcgga tgaatggcag aaattcgaaa gcaaattcga
      541 cccggtcgtc ggttcagggc agggtcgtta aatagccgct tatgtctatt gctggtttac
      601 cggtttattg actaccggaa gcagtgtgac cgtgtgcttc tcaaatgcct gaggccagtt
      661 tgctcaggct ctccccgtgg aggtaataat tgacgatatg atcatttatt ctgcctccca
      721 gagcctgata aaaacggtta gcgcttcgtt aatacagatg taggtgttcc acagggtagc
      781 cagcagcatc ctgcgatgca gatccggaac ataatggtgc agggcgcttg tttcggcgtg
      841 ggtatggtgg caggccccgt ggccggggga ctgttgggcg ctgccggcac ctgtcctacg
      901 agttgcatga taaagaagac agtcataagt gcggcgacga tagtcatgcc ccgcgcccac
      961 cggaaggagc taccggacag cggtgcggac tgttgtaact cagaataaga aatgaggccg
     1021 ctcatggcgt tgactctcag tcatagtatc gtggtatcac cggttggttc cactctctgt
     1081 tgcgggcaac ttcagcagca cgtaggggac ttccgcgttt ccagacttta cgaaacacgg
     1141 aaaccgaaga ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt
     1201 cacgttcgct cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct
     1261 agccgggtcc tcaacgacag gagcacgatc atgcgcaccc gtggccagga cccaacgctg
     1321 cccgagatgc gccgcgtgcg gctgctggag atggcggacg cgatggatat gttctgccaa
     1381 gggttggttt gcgcattcac agttctccgc aagaattgat tggctccaat tcttggagtg
     1441 gtgaatccgt tagcgaggtg ccgccggctt ccattcaggt cgaggtggcc cggctccatg
     1501 caccgcgacg caacgcgggg aggcagacaa ggtatagggc ggcgcctaca atccatgcca
     1561 acccgttcca tgtgctcgcc gaggcggcat aaatcgccgt gacgatcagc ggtccagtga
     1621 tcgaagttag gctggtaaga gccgcgagcg atccttgaag ctgtccctga tggtcgtcat
     1681 ctacctgcct ggacagcatg gcctgcaacg cgggcatccc gatgccgccg gaagcgagaa
     1741 gaatcataat ggggaaggcc atccagcctc gcgtcgcgaa cgccagcaag acgtagccca
     1801 gcgcgtcggc cgccatgccg gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg
     1861 gaccagtgac gaaggcttga gcgagggcgt gcaagattcc gaataccgca agcgacaggc
     1921 cgatcatcgt cgcgctccag cgaaagcggt cctcgccgaa aatgacccag agcgctgccg
     1981 gcacctgtcc tacgagttgc atgataaaga agacagtcat aagtgcggcg acgatagtca
     2041 tgccccgcgc ccaccggaag gagctgactg ggttgaaggc tctcaagggc atcggtcgac
     2101 gctctccctt atgcgactcc tgcattagga agcagcccag tagtaggttg aggccgttga
     2161 gcaccgccgc cgcaaggaat ggtgcatgca aggagatggc gcccaacagt cccccggcca
     2221 cggggcctgc caccataccc acgccgaaac aagcgctcat gagcccgaag tggcgagccc
     2281 gatcttcccc atcggtgatg tcggcgatat aggcgccagc aaccgcacct gtggcgccgg
     2341 tgatgccggc cacgatgcgt ccggcgtaga ggatcagact aagatctcga tcccgcgaaa
     2401 ttaatacgac tcactatagg ggaattgtga gcggataaca attccccact agtaataatt
     2461 ttgtttaact ttaagaagga gatataccat ggcccctata ctaggttatt ggaaaattaa
     2521 gggccttgtg caacccactc gacttctttt ggaatatctt gaagaaaaat atgaagagca
     2581 tttgtatgag cgcgatgaag gtgataaatg gcgaaacaaa aagtttgaat tgggtttgga
     2641 gtttcccaat cttccttatt atattgatgg tgatgttaaa ttaacacagt ctatggccat
     2701 catacgttat atagctgaca agcacaacat gttgggtggt tgtccaaaag agcgtgcaga
     2761 gatttcaatg cttgaaggag cggttttgga tattagatac ggtgtttcga gaattgcata
     2821 tagtaaagac tttgaaactc tcaaagttga ttttcttagc aagctacctg aaatgctgaa
     2881 aatgttcgaa gatcgtttat gtcataaaac atatttaaat ggtgatcatg taacccatcc
     2941 tgacttcatg ttgtatgacg ctcttgatgt tgttttatac atggacccaa tgtgcctgga
     3001 tgcgttccca aaattagttt gttttaaaaa acgtattgaa gctatcccac aaattgataa
     3061 gtacttgaaa tccagcaagt atatagcatg gcctttgcag ggctggcaag ccacgtttgg
     3121 tggtggcgac catccgccaa aatcggatgg taccggcagc ggcctggaag ttctgttcca
     3181 ggggccccat atgctcgagt tcgacatgtg gaaggactac ttcaacctga gccaggtcgt
     3241 gtgggccctg ggcagcagct ggagccaccc gcagttcgaa aaataattaa ttctagagct
     3301 agccctagga gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg ctgctgccac
     3361 cgctgagcaa taactagcat aaccccttgg ggcctctaaa cgggtcttga ggggtttttt
     3421 gctgaaagga ggaactatat ccggatatcc cgcaagaggc ccggcagtac cggcataacc
     3481 aagcctatgc ctacagcatc cagggtgacg gtgccgagga tgacgatgag cgcattgtta
     3541 gatttcatac acggtgcctg actgcgttag caatttaact gtgataaact accgcattaa
     3601 agcttatcga tgataagctg tcaaacatga gaattacaac ttatatcgta tggggctgac
     3661 ttcaggtgct acatttgaag agataaattg cactgaaatc aagaaatatt ttatctgatt
     3721 aataagatga tcttcttgag atcgttttgg tctgcgcgta atctcttgct ctgaaaacga
     3781 aaaaaccgcc ttgcagggcg gtttttcgaa ggttctctga gctaccaact ctttgaaccg
     3841 aggtaactgg cttggaggag cgcagtcacc aaaacttgtc ctttcagttt agccttaacc
     3901 ggcgcatgac ttcaagacta actcctctaa atcaattacc agtggctgct gccagtggtg
     3961 cttttgcatg tctttccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     4021 cggactgaac ggggggttcg tgcatacagt ccagcttgga gcgaactgcc tacccggaac
     4081 tgagtgtcag gcgtggaatg agacaaacgc ggccataaca gcggaatgac accggtaaac
     4141 cgaaaggcag gaacaggaga gcgcacgagg gagccgccag ggggaaacgc ctggtatctt
     4201 tatagtcctg tcgggtttcg ccaccactga tttgagcgtc agatttcgtg atgcttgtca
     4261 ggggggcgga gcctatggaa aaacggcttt gccgcggccc tctcacttcc ctgttaagta
     4321 tcttcctggc atcttccagg aaatctccgc cccgttcgta agccatttcc gctcgccgca
     4381 gtcgaacgac cgagcgtagc gagtcagtga gcgaggaagc ggaatatatc ctgtatcaca
     4441 tattctgctg acgcaccggt gcagcctttt ttctcctgcc acatgaagca cttcactgac
     4501 accctcatca gtgccaacat agtaagccag tatacactcc cctagcgctg atgtccggcg
     4561 gtgcttttgc cgttacgcac caccccgtca gtagctgaac aggagggaca gctgatagaa
     4621 acagaagcca ctggagcacc tcaaaaacac catcatacac taaatcagta agttggcagc
     4681 atcacccgac gcactttgcg ccgaataaat acctgtgacg gaagatcact tcgcagaata
     4741 aataaatcct ggtgtccctg ttgataccgg gaagccctgg gccaactttt ggcgaaaatg
     4801 agacgttgat cggcacgtaa gaggttccaa ctttcaccat aatgaaataa gatcactacc
     4861 gggcgtattt tttgagttat cgagattttc aggagctaag gaagctaaaa tggagaaaaa
     4921 aatcactgga tataccaccg ttgatatatc ccaatggcat cgtaaagaac attttgaggc
     4981 atttcagtca gttgctcaat gtacctataa ccagaccgtt cagctggata ttacggcctt
     5041 tttaaagacc gtaaagaaaa ataagcacaa gttttatccg gcctttattc acattcttgc
     5101 ccgcctgatg aatgctcatc cggaattc
//
