LOCUS       sequence-243824-        5827 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian vector for parallel SLIC cloning containing the VEGF 
            signal sequence for secretion, N-terminal His, PreScission Cleavage 
            site and different C-tag options.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5827)
  AUTHORS   Scholz J, Besir H, Strasser C, Suppmann S
  TITLE     A new method to customize protein expression vectors for fast, 
            efficient and background free parallel cloning.
  JOURNAL   BMC Biotechnol. 2013 Feb 14;13(1):12.
  PUBMED    23410102
REFERENCE   2  (bases 1 to 5827)
  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: "BMC 
            Biotechnol."; date: "2013-02-14"; volume: "13(1)"; pages: "12"
COMMENT     Sequence Label: pCoofy57
FEATURES             Location/Qualifiers
     source          1..5827
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..18
                     /label=L4440
                     /note="L4440 vector, forward primer"
     enhancer        65..444
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        445..648
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     599..619
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     645..669
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     primer_bind     1150..1169
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             1327..1350
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     CDS             1357..1380
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     RBS             1417..1425
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     CDS             1427..1735
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MIQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSD
                     KVSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     1614..1633
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1739..1768
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             join(1772..1777,1778..1801,1802..1837,1838..1861)
                     /codon_start=1
                     /product="two Strep-Tag(R)II moieties connected by a linker
                     for enhanced binding to Strep-Tactin(R), an engineered form
                     of streptavidin (Schmidt et al., 2013)"
                     /label=Twin-Strep-tag
                     /translation="SAWSHPQFEKGGGSGGGSGGSAWSHPQFEK"
     CDS             1865..1909
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             1916..1993
                     /codon_start=1
                     /product="calmodulin-binding peptide"
                     /label=CBP
                     /note="derived from skeletal muscle myosin light chain 
                     kinase; binds calmodulin with nanomolar affinity in the 
                     presence of calcium"
                     /translation="KRRWKKNFIAVSAANRFKKISSSGAL"
     CDS             2000..2035
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     polyA_signal    2726..2781
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2780..2799)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3864..3882)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3950..4054
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4055..4123,4124..4915)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4273..4292)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5086..5674
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5575..5594
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agcgagtcag tgagcgagga agcgtacatt tatattggct catgtccaat atgaccgcca
       61 tgttgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
      121 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      181 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      241 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      301 catcaagtgt atcatatgcc aagtccgccc cctattgacg tcaatgacgg taaatggccc
      361 gcctggcatt atgcccagta catgacctta cgggactttc ctacttggca gtacatctac
      421 gtattagtca tcgctattac catggtgatg cggttttggc agtacaccaa tgggcgtgga
      481 tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg
      541 ttttggcacc aaaatcaacg ggactttcca aaatgtcgta ataaccccgc cccgttgacg
      601 caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctcg tttagtgaac
      661 cgtcagatcc tcactctctt ccgcatcgct gtctgcgagg gccagctgtt gggctcgcgg
      721 ttgaggacaa actcttcgcg gtctttccag tactcttgga tcggaaaccc gtcggcctcc
      781 gaacggtact ccgccaccga gggacctgag cgagtccgca tcgaccggat cggaaaacct
      841 ctcgagaaag gcgtctaacc agtcacagtc gcaaggtagg ctgagcaccg tggcgggcgg
      901 cagcgggtgg cggtcggggt tgtttctggc ggaggtgctg ctgatgatgt aattaaagta
      961 ggcggtcttg agacggcgga tggtcgaggt gaggtgtggc aggcttgaga tccagctgtt
     1021 ggggtgagta ctccctctca aaagcgggca ttacttctgc gctaagattg tcagtttcca
     1081 aaaacgagga ggatttgata ttcacctggc ccgatctggc catacacttg agtgacaatg
     1141 acatccactt tgcctttctc tccacaggtg tccactccca ggtccaagtt taaacggatc
     1201 tctagcgaat tcatgaactt tctgctgtcc tgggtgcact ggtccctggc cctgctgctg
     1261 tacctgcacc acgccaagtg gtcccaggcc gctcctatgg ccgagggcgg tggccagaac
     1321 gctagtcatc accatcacca tcaccatcac gctagcctgg aagttctgtt ccaggggccc
     1381 ttgacaacat aaaaactttg tgttatactt gtaacgtaag gaggtaatga ttcagtttaa
     1441 ggtttacacc tataaaagag agagccgtta tcgtctgttt gtggatgtac agagtgatat
     1501 tattgacacg cccgggcgac ggatggtgat ccccctggcc agtgcacgtc tgctgtcaga
     1561 taaagtctcc cgtgaacttt acccggtggt gcatatcggg gatgaaagct ggcgcatgat
     1621 gaccaccgat atggccagtg tgccggtctc cgttatcggg gaagaagtgg ctgatctcag
     1681 ccaccgcgaa aatgacatca aaaacgccat taacctgatg ttctggggaa tataagagca
     1741 tcatcatcat caccaccacc accaccactg aagcgcttgg agccacccgc agttcgaaaa
     1801 aggtggaggt tctggcggtg gatcgggagg ttcagcgtgg agccacccgc agttcgagaa
     1861 atagaaagaa accgctgctg ctaaattcga acgccagcac atggacagct agatgaagcg
     1921 gcggtggaag aaaaacttta tcgccgtgtc cgccgccaac cggttcaaga agatcagcag
     1981 ctctggcgcc ctgctgtgag aggaccaggt ggacccccgg ctgatcgatg gcaaatgagg
     2041 cagcggcaag atcctgcaca accagaacgt gaacagctgg ggccccatca ccgtgacccc
     2101 taccaccgac ggcggcgaga caagattcga cggccagatc atcgtgcaga tggaaaacga
     2161 ccccgtggtg gccaaggccg ctgccaatct ggccggcaag cacgccgaga gcagcgtggt
     2221 ggtgcagctg gacagcgacg gcaactaccg ggtggtgtac ggcgacccca gcaagctgga
     2281 cggcaagctg cggtggcagc tggtgggaca cggcagggac cacagcgaga caaacaacac
     2341 ccggctgagc ggctacagcg ccgacgagct ggccgtgaag ctggccaagt tccagcagag
     2401 cttcaaccag gccgagaaca tcaacaacaa gcccgaccac atcagcatcg tgggctgcag
     2461 cctggtgtcc gacgacaagc agaagggctt cggccaccag ttcatcaacg ctatggacgc
     2521 caacggcctg cgggtggacg tgtccgtgcg gtctagcgaa ctggccgtgg acgaggccgg
     2581 cagaaagcac accaaggacg ccaatggcga ctgggtgcag aaggccgaaa acaacaaggt
     2641 gtccctgagc tgggacgccc agggcctgga atgagcggcc gctcgaggcc ggcaaggccg
     2701 gatcccccga cctcgacctc tggctaataa aggaaattta ttttcattgc aatagtgtgt
     2761 tggaattttt tgtgtctctc actcggaagg acatatggga gggcaaatca tttggtcgag
     2821 atccctcgga gatctctagc tagaggatcg atccccgccc cggacgaact aaacctgact
     2881 acgacatctc tgccccttct tcgcggggca gtgcatgtaa tcccttcagt tggttggtac
     2941 aacttgccaa ctgaacccta aacgggtagc atatgcttcc cgggtagtag tatatactat
     3001 ccagactaac cctaattcaa tagcatatgt tacccaacgg gaagcatatg ctatcgaatt
     3061 agggttagta aaagggtcct aaggaacagc gatgtaggtg ggcgggccaa gataggggcg
     3121 cgattgctgc gatctggagg acaaattaca cacacttgcg cctgagcgcc aagcacaggg
     3181 ttgttggtcc tcatattcac gaggtcgctg agagcacggt gggctaatgt tgccatgggt
     3241 agcatatact acccaaatat ctggatagca tatgctatcc taatctatat ctgggtagca
     3301 taggctatcc taatctatat ctgggtagca tatgctatcc taatctatat ctgggtagta
     3361 tatgctatcc taatttatat ctgggtagca taggctatcc taatctatat ctgggtagca
     3421 tatgctatcc taatctatat ctgggtagta tatgctatcc taatctgtat ccgggtagca
     3481 tatgctatcc taatagagat tagggtagta tatgctatcc taatttatat ctgggtagca
     3541 tatactaccc aaatatctgg atagcatatg ctatcctaat ctatatctgg gtagcatatg
     3601 ctatcctaat ctatatctgg gtagcatagg ctatcctaat ctatatctgg gtagcatatg
     3661 ctatcctaat ctatatctgg gtagtatatg ctatcctaat ttatatctgg gtagcatagg
     3721 ctatcctaat ctatatctgg gtagcatatg ctatcctaat ctatatctgg gtagtatatg
     3781 ctatcctaat ctgtatccgg gtagcatatg ctatcctcat gataagctgt caaacatgag
     3841 aattaattct tgaagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat
     3901 gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     3961 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     4021 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     4081 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     4141 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     4201 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     4261 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtgtt gacgccgggc aagagcaact
     4321 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     4381 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     4441 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     4501 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     4561 agccatacca aacgacgagc gtgacaccac gatgcctgca gcaatggcaa caacgttgcg
     4621 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     4681 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     4741 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc
     4801 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     4861 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     4921 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     4981 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     5041 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     5101 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     5161 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     5221 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     5281 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     5341 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     5401 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     5461 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     5521 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     5581 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     5641 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     5701 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     5761 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac
     5821 cgagcgc
//
