LOCUS       sequence-361392-        5522 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pET based vector for protein expression in E.coli for targets fused 
            with C-terminal STREP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5522)
  AUTHORS   Bruni R
  TITLE     Bruni Lab Empty Backbones
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5522)
  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"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pC23-LIC-STREP
FEATURES             Location/Qualifiers
     source          1..5522
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      43..498
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(130..149)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     340..361
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        524..628
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(629..697,698..1489)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(847..866)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1660..2248
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2149..2168
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2402..2419
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2434..2573
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2499..2518)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2659..2681
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2675..2866)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     3375..3394
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3375..3393
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(3885..3971,3972..5306)
                     /codon_start=1
                     /gene="Bacillus subtilis sacB"
                     /product="secreted levansucrase that renders bacterial 
                     growth sensitive to sucrose"
                     /label=SacB
                     /note="negative selection marker"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYKVPEFDSSTIKNISSAKGLDVWDSWPLQNTDGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     CDS             5343..5363
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             5364..5387
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             5397..5414
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5467..5485)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      join(5481..5522,1..6)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 tttttgctga aaggaggaac tatatccgga ttggcgaatg ggacgcgccc tgtagcggcg
       61 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc
      121 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
      181 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
      241 accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
      301 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
      361 gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt
      421 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa
      481 tattaacgct tacaatttag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
      541 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
      601 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
      661 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
      721 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
      781 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
      841 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
      901 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
      961 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     1021 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     1081 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     1141 accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt tgcgcaaact
     1201 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     1261 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     1321 taaatctgga gccggtgagc gtggatctcg cggtatcatt gcagcactgg ggccagatgg
     1381 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     1441 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     1501 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     1561 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     1621 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     1681 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     1741 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     1801 tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     1861 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     1921 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     1981 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     2041 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     2101 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     2161 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     2221 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     2281 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     2341 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     2401 cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca
     2461 tctgtgcggt atttcacacc gcaatggtgc actctcagta caatctgctc tgatgccgca
     2521 tagttaagcc agtatacact ccgctatcgc tacgtgactg ggtcatggct gcgccccgac
     2581 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     2641 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     2701 aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc gtgaagcgat tcacagatgt
     2761 ctgcctgttc atccgcgtcc agctcgttga gtttctccag aagcgttaat gtctggcttc
     2821 tgataaagcg ggccatgtta agggcggttt tttcctgttt ggtcactgat gcctccgtgt
     2881 aagggggatt tctgttcatg ggggtaatga taccgatgaa acgagagagg atgctcacga
     2941 tacgggttac tgatgatgaa catgcccggt tactggaacg ttgtgagggt aaacaactgg
     3001 cggtatggat gcggcgggac cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta
     3061 atacagatgt aggtgttcca cagggtagcc agcagcatcc tgcgatgcag atccggaaca
     3121 taatggtgca gggcgctgac ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga
     3181 ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct
     3241 cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct agccgggtcc
     3301 tcaacgacag gagcacgatc atgcgcaccc gtggccagga cccaacgctg cccgagatct
     3361 cgatcccgcg aaattaatac gactcactat agggagtcca caacggtttc cctctagacc
     3421 ttaagaagga gatatactat gcaggtcgtt cactattatt atatttagtg aaatgagata
     3481 ttatgatatt ttctgaattg tgattaaaaa ggcaacttta tgcccatgca acagaaacta
     3541 taaaaaatac agagaatgaa aagaaacaga tagatttttt agttctttag gcccgtagtc
     3601 tgcaaatcct tttatgattt tctatcaaac aaaagaggaa aatagaccag ttgcaatcca
     3661 aacgagagtc taatagaatg aggtcgaaaa gtaaatcgcg cgggtttgtt actgataaag
     3721 caggcaagac ctaaaatgtg taaagggcaa agtgtatact ttggcgtcac cccttacata
     3781 ttttaggtct ttttttattg tgcgtaacta acttgccatc ttcaaacagg agggctggaa
     3841 gaagcagacc gctaacacag tacataaaaa aggagacatg aacgatgaac atcaaaaagt
     3901 ttgcaaaaca agcaacagta ttaaccttta ctaccgcact gctggcagga ggcgcaactc
     3961 aagcgtttgc gaaagaaacg aaccaaaagc catataagga aacatacggc atttcccata
     4021 ttacacgcca tgatatgctg caaatccctg aacagcaaaa aaatgaaaaa tataaagttc
     4081 ctgagttcga ttcgtccaca attaaaaata tctcttctgc aaaaggcctg gacgtttggg
     4141 acagctggcc attacaaaac actgacggca ctgtcgcaaa ctatcacggc taccacatcg
     4201 tctttgcatt agccggagat cctaaaaatg cggatgacac atcgatttac atgttctatc
     4261 aaaaagtcgg cgaaacttct attgacagct ggaaaaacgc tggccgcgtc tttaaagaca
     4321 gcgacaaatt cgatgcaaat gattctatcc taaaagacca aacacaagaa tggtcaggtt
     4381 cagccacatt tacatctgac ggaaaaatcc gtttattcta cactgatttc tccggtaaac
     4441 attacggcaa acaaacactg acaactgcac aagttaacgt atcagcatca gacagctctt
     4501 tgaacatcaa cggtgtagag gattataaat caatctttga cggtgacgga aaaacgtatc
     4561 aaaatgtaca gcagttcatc gatgaaggca actacagctc aggcgacaac catacgctga
     4621 gagatcctca ctacgtagaa gataaaggcc acaaatactt agtatttgaa gcaaacactg
     4681 gaactgaaga tggctaccaa ggcgaagaat ctttatttaa caaagcatac tatggcaaaa
     4741 gcacatcatt cttccgtcaa gaaagtcaaa aacttctgca aagcgataaa aaacgcacgg
     4801 ctgagttagc aaacggcgct ctcggtatga ttgagctaaa cgatgattac acactgaaaa
     4861 aagtgatgaa accgctgatt gcatctaaca cagtaacaga tgaaattgaa cgcgcgaacg
     4921 tctttaaaat gaacggcaaa tggtacctgt tcactgactc ccgcggatca aaaatgacga
     4981 ttgacggcat tacgtctaac gatatttaca tgcttggtta tgtttctaat tctttaactg
     5041 gcccatacaa gccgctgaac aaaactggcc ttgtgttaaa aatggatctt gatcctaacg
     5101 atgtaacctt tacttactca cacttcgctg tacctcaagc gaaaggaaac aatgtcgtga
     5161 ttacaagcta tatgacaaac agaggattct acgcagacaa acaatcaacg tttgcgccta
     5221 gcttcctgct gaacatcaaa ggcaagaaaa catctgttgt caaagacagc atccttgaac
     5281 aaggacaatt aacagttaac aaataaaaac gcaaaagaaa attttagtaa gccacctgcg
     5341 ccgaaaacct ctattttcag ggatggagcc acccgcagtt cgaaaaataa ctcgagcacc
     5401 accaccacca ccactgagat ccggctgcta acaaagcccg aaaggaagct gagttggctg
     5461 ctgccaccgc tgagcaataa ctagcataac cccttggggc ctctaaacgg gtcttgaggg
     5521 gt
//
