LOCUS       sequence-212122-        6829 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway destination vector .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6829)
  AUTHORS   Lu G, Middleton RE, Sun H, Naniong M, Ott CJ, Mitsiades CS, Wong KK,
            Bradner JE, Kaelin WG Jr
  TITLE     The myeloma drug lenalidomide promotes the cereblon-dependent 
            destruction of Ikaros proteins.
  JOURNAL   Science. 2014 Jan 17;343(6168):305-9. doi: 10.1126/science.1244917. 
            Epub 2013 Nov 29.
  PUBMED    24292623
REFERENCE   2  (bases 1 to 6829)
  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: 
            "10.1126/science.1244917"; journalName: "Science"; date: 
            "2014-01-17- 17"; volume: "343"; issue: "6168"; pages: "305-9"
COMMENT     Sequence Label: pcDNA3-FLAG- gate-pGK-HYG
FEATURES             Location/Qualifiers
     source          1..6829
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        190..569
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        570..773
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     724..744
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     818..837
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        818..836
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      872..881
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             881..904
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     protein_bind    938..1062
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(1087..1092,1093..1110,1111..1117)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             1171..1851
                     /codon_start=1
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQAGRNLE
                     DPAY"
     primer_bind     complement(1237..1256)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             2171..2476
                     /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
                     VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     2355..2374
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(2517..2641)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        2785..3284
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2820..2843)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3217..3236
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3302..4327
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             3302..4327
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    4457..4578
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4494..4513)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4548..4567
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4627..4643)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4627..4643)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4640..4662)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4651..4667
                     /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(4675..4681,4682..4699,4700..4705))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4720..4741
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4858..4875)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5029..5617)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5109..5128)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5788..6579,6580..6648))
                     /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     6411..6430
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6649..6753)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(join(6828..6829,1..18))
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 caatctgctc tgatgccgca tagttaagcc agtatctgct ccctgcttgt gtgttggagg
       61 tcgctgagta gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg accgacaatt
      121 gcatgaagaa tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat
      181 atacgcgttg acattgatta ttgactagtt attaatagta atcaattacg gggtcattag
      241 ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct
      301 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc
      361 caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg
      421 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
      481 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
      541 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
      601 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
      661 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
      721 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctctggc
      781 taactagaga acccactgct tactggctta tcgaaattaa tacgactcac tatagggaga
      841 cccaagctgg ctagcgttta aacttaagct tgccaccatg gactacaagg acgacgatga
      901 caagctcgat ggaggtgcgg ccgctcgagg ttatcaaaca agtttgtaca aaaaagctga
      961 acgagaaacg taaaatgata taaatatcaa tatattaaat tagattttgc ataaaaaaca
     1021 gactacataa tactgtaaaa cacaacatat ccagtcacta tggcggccgc attaggcacc
     1081 ccaggcttta cactttatgc ttccggctcg tataatgtgt ggattttgag ttaggatccg
     1141 tcgagatttt caggagctaa ggaagctaaa atggagaaaa aaatcactgg atataccacc
     1201 gttgatatat cccaatggca tcgtaaagaa cattttgagg catttcagtc agttgctcaa
     1261 tgtacctata accagaccgt tcagctggat attacggcct ttttaaagac cgtaaagaaa
     1321 aataagcaca agttttatcc ggcctttatt cacattcttg cccgcctgat gaatgctcat
     1381 ccggaattcc gtatggcaat gaaagacggt gagctggtga tatgggatag tgttcaccct
     1441 tgttacaccg ttttccatga gcaaactgaa acgttttcat cgctctggag tgaataccac
     1501 gacgatttcc ggcagtttct acacatatat tcgcaagatg tggcgtgtta cggtgaaaac
     1561 ctggcctatt tccctaaagg gtttattgag aatatgtttt tcgtctcagc caatccctgg
     1621 gtgagtttca ccagttttga tttaaacgtg gccaatatgg acaacttctt cgcccccgtt
     1681 ttcaccatgg gcaaatatta tacgcaaggc gacaaggtgc tgatgccgct ggcgattcag
     1741 gttcatcatg ccgtttgtga tggcttccat gtcggcagaa tgcttaatga attacaacag
     1801 tactgcgatg agtggcaggc ggggcgtaat ctagaggatc cggcttacta aaagccagat
     1861 aacagtatgc gtatttgcgc gctgattttt gcggtataag aatatatact gatatgtata
     1921 cccgaagtat gtcaaaaaga ggtatgctat gaagcagcgt attacagtga cagttgacag
     1981 cgacagctat cagttgctca aggcatatat gatgtcaata tctccggtct ggtaagcaca
     2041 accatgcaga atgaagcccg tcgtctgcgt gccgaacgct ggaaagcgga aaatcaggaa
     2101 gggatggctg aggtcgcccg gtttattgaa atgaacggct cttttgctga cgagaacagg
     2161 ggctggtgaa atgcagttta aggtttacac ctataaaaga gagagccgtt atcgtctgtt
     2221 tgtggatgta cagagtgata ttattgacac gcccgggcga cggatggtga tccccctggc
     2281 cagtgcacgt ctgctgtcag ataaagtccc ccgtgaactt tacccggtgg tgcatatcgg
     2341 ggatgaaagc tggcgcatga tgaccaccga tatggccagt gtgccggtct ccgttatcgg
     2401 ggaagaagtg gctgatctca gccaccgcga aaatgacatc aaaaacgcca ttaacctgat
     2461 gttctgggga atataaatgt caggctccct tatacacagc cagtctgcag gtcgaccata
     2521 gtgactggat atgttgtgtt ttacagtatt atgtagtctg ttttttatgc aaaatctaat
     2581 ttaatatatt gatatttata tcattttacg tttctcgttc agctttcttg tacaaagtgg
     2641 ttcgataacg aattccgccc ccccccccta acgttactgg ccgaagccgc ttggaataag
     2701 gccggtgtgc gtttgtctat atgttatttt ccaccatatt gccgtctttt ggcaatgtga
     2761 gggccgcgtt taacgaattc taccgggtag gggaggcgct tttcccaagg cagtctggag
     2821 catgcgcttt agcagccccg ctgggcactt ggcgctacac aagtggcctc tggcctcgca
     2881 cacattccac atccaccggt aggcgccaac cggctccgtt ctttggtggc cccttcgcgc
     2941 caccttctac tcctccccta gtcaggaagt tcccccccgc cccgcagctc gcgtcgtgca
     3001 ggacgtgaca aatggaagta gcacgtctca ctagtctcgt gcagatggac agcaccgctg
     3061 agcaatggaa gcgggtaggc ctttggggca gcggccaata gcagctttgc tccttcgctt
     3121 tctgggctca gaggctggga aggggtgggt ccgggggcgg gctcaggggc gggctcaggg
     3181 gcggggcggg cgcccgaagg tcctccggag gcccggcatt ctgcacgctt caaaagcgca
     3241 cgtctgccgc gctgttctcc tcttcctcat ctccgggcct ttcgacctgc atcccgccac
     3301 catgaaaaag cctgaactca ccgcgacgtc tgtcgagaag tttctgatcg aaaagttcga
     3361 cagcgtctcc gacctgatgc agctctcgga gggcgaagaa tctcgtgctt tcagcttcga
     3421 tgtaggaggg cgtggatatg tcctgcgggt aaatagctgc gccgatggtt tctacaaaga
     3481 tcgttatgtt tatcggcact ttgcatcggc cgcgctcccg attccggaag tgcttgacat
     3541 tggggaattc agcgagagcc tgacctattg catctcccgc cgtgcacagg gtgtcacgtt
     3601 gcaagacctg cctgaaaccg aactgcccgc tgttctgcag ccggtcgcgg aggccatgga
     3661 tgcgatcgct gcggccgatc ttagccagac gagcgggttc ggcccattcg gaccgcaagg
     3721 aatcggtcaa tacactacat ggcgtgattt catatgcgcg attgctgatc cccatgtgta
     3781 tcactggcaa actgtgatgg acgacaccgt cagtgcgtcc gtcgcgcagg ctctcgatga
     3841 gctgatgctt tgggccgagg actgccccga agtccggcac ctcgtgcacg cggatttcgg
     3901 ctccaacaat gtcctgacgg acaatggccg cataacagcg gtcattgact ggagcgaggc
     3961 gatgttcggg gattcccaat acgaggtcgc caacatcttc ttctggaggc cgtggttggc
     4021 ttgtatggag cagcagacgc gctacttcga gcggaggcat ccggagcttg caggatcgcc
     4081 gcggctccgg gcgtatatgc tccgcattgg tcttgaccaa ctctatcaga gcttggttga
     4141 cggcaatttc gatgatgcag cttgggcgca gggtcgatgc gacgcaatcg tccgatccgg
     4201 agccgggact gtcgggcgta cacaaatcgc ccgcagaagc gcggccgtct ggaccgatgg
     4261 ctgtgtagaa gtactcgccg atagtggaaa ccgacgcccc agcactcgtc cgagggcaaa
     4321 ggaatagcac gtactacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     4381 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     4441 gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     4501 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     4561 actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta
     4621 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     4681 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     4741 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     4801 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     4861 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     4921 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     4981 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     5041 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     5101 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     5161 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     5221 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     5281 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     5341 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     5401 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     5461 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     5521 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     5581 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     5641 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     5701 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     5761 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     5821 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     5881 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     5941 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     6001 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     6061 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     6121 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     6181 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     6241 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     6301 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     6361 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     6421 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     6481 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     6541 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     6601 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     6661 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     6721 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     6781 cgtcgacgga tcgggagatc tcccgatccc ctatggtgca ctctcagta
//
