LOCUS       sequence-248426-        5932 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A pCS2 based vector to generate a 3'-Flag tag using GATEWAY cloning 
            technology.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5932)
  AUTHORS   Grant IM, Balcha D, Hao T, Shen Y, Trivedi P, Patrushev I, Fortriede
            JD, Karpinka JB, Liu L, Zorn AM, Stukenberg PT, Hill DE, Gilchrist 
            MJ
  TITLE     The Xenopus ORFeome: A resource that enables functional genomics.
  JOURNAL   Dev Biol. 2015 Dec 15;408(2):345-57. doi: 
            10.1016/j.ydbio.2015.09.004. Epub 2015 Sep 29.
  PUBMED    26391338
REFERENCE   2  (bases 1 to 5932)
  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: 
            "10.1016/j.ydbio.2015.09.004"; journalName: "Dev Biol"; date: 
            "2015-12-15- 15"; volume: "408"; issue: "2"; pages: "345-57"
COMMENT     Sequence Label: pCSF107mT-GATEWAY-3'-FLAG
FEATURES             Location/Qualifiers
     source          1..5932
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..978
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     promoter        1013..1031
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1013..1030
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     protein_bind    1096..1220
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(1245..1250,1251..1268,1269..1275)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             1329..1988
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1395..1414)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             2309..2614
                     /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     2493..2512
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(2655..2779)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     regulatory      2787..2796
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2796..2861
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(2896..2915)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2897..2915)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    2920..3054
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2944..2963)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2998..3017
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(3205..3225)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3205..3223)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3244..3260)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3244..3260)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3257..3279)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3268..3284
                     /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)."
     promoter        complement(join(3292..3298,3299..3316,3317..3322))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3337..3358
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3475..3492)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3646..4234)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3726..3745)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4405..5196,5197..5265))
                     /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     5028..5047
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5266..5370)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5396..5851)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5533..5554)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5745..5764
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gccaattcaa tatggcgtat atggactcat gccaattcaa tatggtggat ctggacctgt
       61 gccaattcaa tatggcgtat atggactcgt gccaattcaa tatggtggat ctggacccca
      121 gccaattcaa tatggcggac ttggcaccat gccaattcaa tatggcggac ctggcactgt
      181 gccaactggg gaggggtcta cttggcacgg tgccaagttt gaggaggggt cttggccctg
      241 tgccaagtcc gccatattga attggcatgg tgccaataat ggcggccata ttggctatat
      301 gccaggatca atatataggc aatatccaat atggccctat gccaatatgg ctattggcca
      361 ggttcaatac tatgtattgg ccctatgcca tatagtattc catatatggg ttttcctatt
      421 gacgtagata gcccctccca atgggcggtc ccatatacca tatatggggc ttcctaatac
      481 cgcccatagc cactccccca ttgacgtcaa tggtctctat atatggtctt tcctattgac
      541 gtcatatggg cggtcctatt gacgtatatg gcgcctcccc cattgacgtc aattacggta
      601 aatggcccgc ctggctcaat gcccattgac gtcaatagga ccacccacca ttgacgtcaa
      661 tgggatggct cattgcccat tcatatccgt tctcacgccc cctattgacg tcaatgacgg
      721 taaatggccc acttggcagt acatcaatat ctattaatag taacttggca agtacattac
      781 tattggaagt acgccagggt acattggcag tactcccatt gacgtcaatg gcggtaaatg
      841 gcccgcgatg gctgccaagt acatccccat tgacgtcaat ggggaggggc aatgacgcaa
      901 atgggcgttc cattgacgta aatgggcggt aggcgtgcct aatgggaggt ctatataagc
      961 aatgctcgtt tagggaaccg ccattctgcc tggggacgtc ggagcaagct tgatttaggt
     1021 gacactatag aatacaagct acttgttctt tttgcaggat ccatcgattt cgaagaattc
     1081 agatctcttg gtaccacaag tttgtacaaa aaagctgaac gagaaacgta aaatgatata
     1141 aatatcaata tattaaatta gattttgcat aaaaaacaga ctacataata ctgtaaaaca
     1201 caacatatcc agtcactatg gcggccgcat taggcacccc aggctttaca ctttatgctt
     1261 ccggctcgta taatgtgtgg attttgagtt aggatccgtc gagattttca ggagctaagg
     1321 aagctaaaat ggagaaaaaa atcactggat ataccaccgt tgatatatcc caatggcatc
     1381 gtaaagaaca ttttgaggca tttcagtcag ttgctcaatg tacctataac cagaccgttc
     1441 agctggatat tacggccttt ttaaagaccg taaagaaaaa taagcacaag ttttatccgg
     1501 cctttattca cattcttgcc cgcctgatga atgctcatcc ggaattccgt atggcaatga
     1561 aagacggtga gctggtgata tgggatagtg ttcacccttg ttacaccgtt ttccatgagc
     1621 aaactgaaac gttttcatcg ctctggagtg aataccacga cgatttccgg cagtttctac
     1681 acatatattc gcaagatgtg gcgtgttacg gtgaaaacct ggcctatttc cctaaagggt
     1741 ttattgagaa tatgtttttc gtctcagcca atccctgggt gagtttcacc agttttgatt
     1801 taaacgtggc caatatggac aacttcttcg cccccgtttt caccatgggc aaatattata
     1861 cgcaaggcga caaggtgctg atgccgctgg cgattcaggt tcatcatgcc gtttgtgatg
     1921 gcttccatgt cggcagaatg cttaatgaat tacaacagta ctgcgatgag tggcagggcg
     1981 gggcgtaaac gcgtggatcc ggcttactaa aagccagata acagtatgcg tatttgcgcg
     2041 ctcgcgaacc ggtgtatacc cgaagtatgt caaaaagagg tatgctatga agcagcgtat
     2101 tacagtgaca gttgacagcg acagctatca gttgctcaag gcatatatga tgtcaatatc
     2161 tccggtctgg taagcacaac catgcagaat gaagcccgtc gtctgcgtgc cgaacgctgg
     2221 aaagcggaaa atcaggaagg gatggctgag gtcgcccggt ttattgaaat gaacggctct
     2281 tttgctgacg agaacagggg ctggtgaaat gcagtttaag gtttacacct ataaaagaga
     2341 gagccgttat cgtctgtttg tggatgtaca gagtgatatt attgacacgc ccgggcgacg
     2401 gatggtgatc cccctggcca gtgcacgtct gctgtcagat aaagtctccc gtgaacttta
     2461 cccggtggtg catatcgggg atgaaagctg gcgcatgatg accaccgata tggccagtgt
     2521 gccggtctcc gttatcgggg aagaagtggc tgatctcagc caccgcgaaa atgacatcaa
     2581 aaacgccatt aacctgatgt tctggggaat ataaatgtca ggctccctta tacacagcca
     2641 gtctgcaggt cgaccatagt gactggatat gttgtgtttt acagtattat gtagtctgtt
     2701 ttttatgcaa aatctaattt aatatattga tatttatatc attttacgtt tctcgttcag
     2761 ctttcttgta caaagtggtt ggtaccgcca ccatggacta caaagaccat gacggtgatt
     2821 ataaagatca tgatatcgat tacaaggatg acgatgacaa ggaattctag ctcgagtcta
     2881 gaatcgagcc tctcgcccta tagtgagtcg tattacgtag atccagacat gataagatac
     2941 attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa
     3001 atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac
     3061 aacaattgca ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaattc
     3121 gcggcgcgcc gcggttccat ctgttttctt atctgttctt tcatctgttc ttttatctgt
     3181 ttgtttggta cccagctttt gttcccttta gtgagggtta attgcgcgct tggcgtaatc
     3241 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     3301 agccgggagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     3361 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     3421 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     3481 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     3541 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     3601 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     3661 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     3721 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     3781 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     3841 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     3901 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     3961 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     4021 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     4081 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     4141 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     4201 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     4261 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     4321 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     4381 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     4441 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     4501 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     4561 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     4621 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     4681 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     4741 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     4801 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     4861 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     4921 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     4981 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     5041 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     5101 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     5161 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     5221 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     5281 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     5341 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctaaatt
     5401 gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt
     5461 aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg
     5521 ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc
     5581 aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca
     5641 agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga
     5701 tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa
     5761 ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc
     5821 gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct gcgcaactgt
     5881 tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agtcgatcga cc
//
