LOCUS       sequence-71987-e        6578 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  FX cloning mammalian cell line expression vector with CMV promoter 
            and N-terminal streptavidin binding peptide, Myc tag, EGFP and 3C 
            protease cleavage site .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6578)
  AUTHORS   Geertsma ER, Dutzler R
  TITLE     A versatile and efficient high-throughput cloning tool for 
            structural biology.
  JOURNAL   Biochemistry. 2011 Apr 19;50(15):3272-8. Epub 2011 Mar 25.
  PUBMED    21410291
REFERENCE   2  (bases 1 to 6578)
  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: 
            "Biochemistry."; date: "2011-04-19"; volume: "50(15)"; pages: 
            "3272-8. Epub 2011 Mar 25"
COMMENT     Sequence Label: pcDXNSMG3
FEATURES             Location/Qualifiers
     source          1..6578
                     /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"
     misc_feature    917..1079
                     /label=QBI SP163
                     /note="strong translational enhancer derived from the 5' 
                     UTR of the mouse VEGF gene"
     CDS             1092..1205
                     /codon_start=1
                     /product="streptavidin-binding peptide"
                     /label=SBP
                     /note="selected from a peptide library; binds streptavidin 
                     with nanomolar affinity (Keefe et al., 2001)"
                     /translation="MDEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP"
     CDS             1206..1235
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             join(1242..1244,1245..1247,1248..1958)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1242..1244,1245..1247,1248..1958)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1287..1308)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1548..1567)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1895..1916
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1965..1988
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             complement(2037..2342)
                     /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="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(2139..2158)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(2684..3343)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     3258..3277
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(3397..3403,3404..3421,3422..3427))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(3452..3576)
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(3595..3615)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(3619..3639)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(3706..3723)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3712..3936
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3982..4410
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4069..4088)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4279..4300
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4419..4439)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4424..4753
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4604..4739
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4666..4685
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     rep_origin      complement(4826..5411)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4906..4925)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5582..6373,6374..6442))
                     /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     6205..6224
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6443..6547)
                     /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 gctggctagc
      901 gtttaaactt aagcttagcg cagaggcttg gggcagccga gcggcagcca ggccccggcc
      961 cgggcctcgg ttccagaagg gagaggagcc cgccaaggcg cgcaagagag cgggctgcct
     1021 cgcagtccga gccggagagg gagcgcgagc cgcgccggcc ccggacggcc tccgaaacca
     1081 tgggggccag catggacgag aagaccactg gttggcgagg tggacacgtt gttgaaggac
     1141 tggctgggga acttgaacaa cttcgtgcac gactggagca tcacccacaa ggtcaacgtg
     1201 aaccagaaca aaaactcatc tcagaagagg atctgggtac catggtgagc aagggcgagg
     1261 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca
     1321 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt
     1381 tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct
     1441 acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt
     1501 ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact
     1561 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga
     1621 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca
     1681 acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca
     1741 agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca
     1801 cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg
     1861 ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg
     1921 ccgccgggat cactctcggc atggacgagc tgtacaaggg taccttagaa gttttgtttc
     1981 aaggtccaag tagaagagcg agctgcagac tggctgtgta taagggagcc tgacatttat
     2041 attccccaga acatcaggtt aatggcgttt ttgatgtcat tttcgcggtg gctgagatca
     2101 gccacttctt ccccgataac ggagaccggc acactggcca tatcggtggt catcatgcgc
     2161 cagctttcat ccccgatatg caccaccggg taaagttcac gggagacttt atctgacagc
     2221 agacgtgcac tggccagggg gatcaccatc cgtcgcccgg gcgtgtcaat aatatcactc
     2281 tgtacatcca caaacagacg ataacggctc tctcttttat aggtgtaaac cttaaactgc
     2341 atttcaccag cccctgttct cgtcagcaaa agagccgttc atttcaataa accgggcgac
     2401 ctcagccatc ccttcctgat tttccgcttt ccagcgttcg gcacgcagac gacgggcttc
     2461 attctgcatg gttgtgctta ccagaccgga gatattgaca tcatatatgc cttgagcaac
     2521 tgatagctgt cgctgtcaac tgtcactgta atacgctgct tcatagcata cctctttttg
     2581 acatacttcg ggtatacata tcagtatata ttcttatacc gcaaaaatca gcgcgcaaat
     2641 acgcatactg ttatctggct tttagtaagc cggatccacg cgtttacgcc ccgccctgcc
     2701 actcatcgca gtactgttgt aattcattaa gcattctgcc gacatggaag ccatcacaaa
     2761 cggcatgatg aacctgaatc gccagcggca tcagcacctt gtcgccttgc gtataatatt
     2821 tgcccatggt gaaaacgggg gcgaagaagt tgtccatatt ggccacgttt aaatcaaaac
     2881 tggtgaaact cacccaggga ttggctgaga cgaaaaacat attctcaata aaccctttag
     2941 ggaaataggc caggttttca ccgtaacacg ccacatcttg cgaatatatg tgtagaaact
     3001 gccggaaatc gtcgtggtat tcactccaga gcgatgaaaa cgtttcagtt tgctcatgga
     3061 aaacggtgta acaagggtga acactatccc atatcaccag ctcaccgtct ttcattgcca
     3121 tacggaattc cggatgagca ttcatcaggc gggcaagaat gtgaataaag gccggataaa
     3181 acttgtgctt atttttcttt acggtcttta aaaaggccgt aatatccagc tgaacggtct
     3241 ggttataggt acattgagca actgactgaa atgcctcaaa atgttcttta cgatgccatt
     3301 gggatatatc aacggtggta tatccagtga tttttttctc cattttagct tccttagctc
     3361 ctgaaaatct cgacggatcc taactcaaaa tccacacatt atacgagccg gaagcataaa
     3421 gtgtaaagcc tggggtgcct aatgcggccg ccatagtgac tggatatgtt gtgttttaca
     3481 gtattatgta gtctgttttt tatgcaaaat ctaatttaat atattgatat ttatatcatt
     3541 ttacgtttct cgttcagctt ttttgtacaa acttgtgata tcgaattcgc tagctacctt
     3601 tctcttcttt tttggatcta cctttctctt cttttttggg gatccactag ttcagctctt
     3661 ctgcataata actatctaga gggcccgttt aaacccgctg atcagcctcg actgtgcctt
     3721 ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg
     3781 ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt
     3841 gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca
     3901 atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct
     3961 ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
     4021 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt
     4081 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc
     4141 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg
     4201 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
     4261 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct
     4321 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg
     4381 agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg
     4441 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     4501 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     4561 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     4621 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     4681 cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     4741 aggcttttgc aaaaagctcc cgggacatgt gagcaaaagg ccagcaaaag gccaggaacc
     4801 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     4861 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     4921 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     4981 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     5041 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     5101 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     5161 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     5221 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     5281 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     5341 aacaaaccac cgctggtagc ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     5401 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     5461 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     5521 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     5581 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     5641 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     5701 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     5761 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     5821 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     5881 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     5941 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     6001 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     6061 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     6121 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     6181 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     6241 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     6301 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     6361 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     6421 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     6481 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     6541 ttccgcgcac atttccccga aaagtgccac ctgacgtc
//
