LOCUS       sequence-324345-        2529 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BB1 encoding a MS2 binding VP64 fusion protein with an additional 
            SV40 NLS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2529)
  AUTHORS   Baumschabl M, Prielhofer R, Mattanovich D, Steiger MG
  TITLE     Fine-Tuning of Transcription in Pichia pastoris Using dCas9 and RNA 
            Scaffolds.
  JOURNAL   ACS Synth Biol. 2020 Dec 18;9(12):3202-3209. doi: 
            10.1021/acssynbio.0c00214. Epub 2020 Nov 12.
  PUBMED    33180466
REFERENCE   2  (bases 1 to 2529)
  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: 
            "10.1021/acssynbio.0c00214"; journalName: "ACS Synth Biol"; date: 
            "2020-12-18- 18"; volume: "9"; issue: "12"; pages: "3202-3209"
COMMENT     Sequence Label: BB1_23_MS2bind_VP64_SV40NLS
FEATURES             Location/Qualifiers
     source          1..2529
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     610..629
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(703..805)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     terminator      complement(806..837)
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     primer_bind     885..901
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     885..901
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             988..1008
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1018..1404
                     /codon_start=1
                     /product="bacteriophage MS2 coat protein"
                     /label=MS2
                     /note="binds to a specific stem-loop structure in the viral
                     RNA (Peabody, 1993)"
                     /translation="ASNFTQFVLVDNGGTGDVTVAPSNFANGVAEWISSNSRSQAYKVT
                     CSVRQSSAQNRKYTIKVEVPKVATQTVGGVELPVAAWRSYLNMELTIPIFATNSDCELI
                     VKAMQGLLKDGNPIPSAIAANSGIY"
     CDS             1420..1569
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     primer_bind     complement(1709..1726)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1709..1725)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      1766..2353
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2254..2273
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(2365..2392)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(join(2416..2529,1..702))
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCLKPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNGDRVFRLAQAQSRMNNGLVGA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
ORIGIN
        1 gttccatagg atggcaagat cctggtatcg gtctgcgatt ccgactcgtc caacatcaat
       61 acaacctatt aatttcccct cgtcaaaaat aaggttatca agtgagaaat caccatgagt
      121 gacgactgaa tccggtgaga atggcaagag tttatgcatt tctttccaga cttgttcaac
      181 aggccagcca ttacgctcgt catcaaaatc actcgcacca accaaaccgt tattcattcg
      241 tgattgcgcc tgagcgagac gaaatacgcg atcgccgtta aaaggacaat tacaaacagg
      301 aatcgaatgc aaccggcgca ggaacactgc cagcgcatca acaatatttt cacctgaatc
      361 aggatattct tctaatacct ggaatgctgt tttccctggg atcgcagtgg tgagtaacca
      421 tgcatcatca ggagtacgga taaaatgctt gatggtcgga agaggcataa attccgtcag
      481 ccagtttagc ctgaccatct catctgtaac atcattggca acgctacctt tgccatgttt
      541 cagaaacaac tctggcgcat cgggcttccc atacaatcga tagattgtcg cacctgattg
      601 cccgacatta tcgcgagccc atttataccc atataaatca gcatccatgt tggaatttaa
      661 tcgcggcttc aagcaagacg tttcccgttg aatatggctc attttagctt ccttagctcc
      721 tgaaaatctc gataactcaa aaaatacgcc cggtagtgat cttatttcat tatggtgaaa
      781 gttggaacct cttacgtgcc gatcaagtca aaagcctccg gtcggaggct tttgactttc
      841 tgctatggag gtcaggtatg atttaaatgg tcagtattga gcctcaggaa acagctatga
      901 catcaagctg actagataat ctagctgatc gtggaccgat catacgtata atgccgtaag
      961 atcacgcgcg aattcgaaga cgccatgccc aaaaagaaaa gaaaagtggg tagtatggcg
     1021 tctaacttca cccagttcgt tctggttgac aacggtggta ccggtgacgt taccgttgcg
     1081 ccgtctaact tcgcgaacgg tgttgcggaa tggatctctt ctaactctcg ttctcaggcg
     1141 tacaaagtta cctgctctgt tcgtcaatcc tccgcgcaga accgtaaata caccatcaaa
     1201 gttgaagttc cgaaagttgc gacccagacc gttggtggtg ttgaactgcc ggttgcggcg
     1261 tggcgttctt acctgaacat ggaactgacc atcccgatct tcgcgaccaa ctctgactgc
     1321 gaactgatcg ttaaagcgat gcagggtctg ctgaaagacg gtaacccgat cccgtctgcg
     1381 atcgcggcga actctggtat ctacggctcc ggacgagccg atgcccttga cgattttgac
     1441 ttggacatgt tgggttctga cgctttggat gactttgatc ttgatatgct tggttccgac
     1501 gctctagatg atttcgactt ggatatgctg ggatccgatg ccttggacga tttcgacttg
     1561 gatatgttga tcaactgagc ttgcgtcttc gaattcgagg ccgcatatca ttcaggacga
     1621 gcctcagact ccagcgtaac tggactgcaa tcaactcact ggctcacctt ccggtccacg
     1681 atcagctaga atcaagctga ctagataaac tggccgtcgt tttacacggg tgggcctttc
     1741 ttcggtagaa aatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     1801 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     1861 ctctttttcc gaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     1921 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     1981 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     2041 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     2101 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     2161 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     2221 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     2281 tgtcgggttt cgccacctct gacttgagca tcgatttttg tgatgctcgt caggggggcg
     2341 gagcctatgg aaaaacgcca gcaacgcaga aaggcccacc cgaaggtgag ccaggtgatt
     2401 acatttaggt cctcattaga aaaactcatc gagcatcaag tgaaactgca atttattcat
     2461 atcaggatta tcaataccat atttttgaaa aagccgtttc tgtaatgaag gagaaaactc
     2521 accgaggca
//
