LOCUS       Exported                7823 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Express  Smad4 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    pLPCX2-FLAG-Smad4
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7823)
  AUTHORS   Sun J, He H, Pillai S, Xiong Y, Challa S, Xu L, Chellappan S, Yang S
  TITLE     GATA3 transcription factor abrogates Smad4 transcription 
            factor-mediated fascin overexpression, invadopodium formation, and 
            breast cancer cell invasion.
  JOURNAL   J Biol Chem. 2013 Dec 27;288(52):36971-82. doi: 
            10.1074/jbc.M113.506535. Epub 2013 Nov 14.
  PUBMED    24235142
REFERENCE   2  (bases 1 to 7823)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7823
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     581..605
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             707..730
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     LTR             2441..3034
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(3137..3159)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3245..3385
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3300..3319
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3400..3417)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3571..4159)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3651..3670)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4330..5190)
                     /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     4953..4972
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5191..5295)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5363..5381
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     LTR             5953..6128
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    6191..6390
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             6591..7007
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     6937..6959
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     6975..6991
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     CDS             7033..7632
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(7033..7052)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7529..7549
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      601 agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagacac cgggaccgat
      661 ccagcctccg cggccccaag ccctcgaggg cttgttcgat accatggact acaaggacga
      721 cgatgacaag ggtcgactgg acaatatgtc tattacgaat acaccaacaa gtaatgatgc
      781 ctgtctgagc attgtgcata gtttgatgtg ccatagacaa ggtggagaga gtgaaacatt
      841 tgcaaaaaga gcaattgaaa gtttggtaaa gaagctgaag gagaaaaaag atgaattgga
      901 ttctttaata acagctataa ctacaaatgg agctcatcct agtaaatgtg ttaccataca
      961 gagaacattg gatgggaggc ttcaggtggc tggtcggaaa ggatttcctc atgtgatcta
     1021 tgcccgtctc tggaggtggc ctgatcttca caaaaatgaa ctaaaacatg ttaaatattg
     1081 tcagtatgcg tttgacttaa aatgtgatag tgtctgtgtg aatccatatc actacgaacg
     1141 agttgtatca cctggaattg atctctcagg attaacactg cagagtaatg ctccatcaag
     1201 tatgatggtg aaggatgaat acgttcatga ctttgaggga cagccatcgt tgtccactga
     1261 aggacattca attcaaacca tccagcatcc accaagtaat cgtgcatcga cagagacata
     1321 cagcacccca gctctgttag ccccatctga gtctaatgct accagcactg ccaactttcc
     1381 caacattcct gtggcttcca caagtcagcc tgccagtata ctggggggca gccatagtga
     1441 aggactgttg cagatagcat cagggcctca gccaggacag cagcagaatg gatttactgg
     1501 tcagccagct acttaccatc ataacagcac taccacctgg actggaagta ggactgcacc
     1561 atacacacct aatttgcctc accaccaaaa cggccatctt cagcaccacc cgcctatgcc
     1621 gccccatccc ggacattact ggcctgttca caatgagctt gcattccagc ctcccatttc
     1681 caatcatcct gctcctgagt attggtgttc cattgcttac tttgaaatgg atgttcaggt
     1741 aggagagaca tttaaggttc cttcaagctg ccctattgtt actgttgatg gatacgtgga
     1801 cccttctgga ggagatcgct tttgtttggg tcaactctcc aatgtccaca ggacagaagc
     1861 cattgagaga gcaaggttgc acataggcaa aggtgtgcag ttggaatgta aaggtgaagg
     1921 tgatgtttgg gtcaggtgcc ttagtgacca cgcggtcttt gtacagagtt actacttaga
     1981 cagagaagct gggcgtgcac ctggagatgc tgttcataag atctacccaa gtgcatatat
     2041 aaaggtcttt gatttgcgtc agtgtcatcg acagatgcag cagcaggcgg ctactgcaca
     2101 agctgcagca gctgcccagg cagcagccgt ggcaggaaac atccctggcc caggatcagt
     2161 aggtggaata gctccagcta tcagtctgtc agctgctgct ggaattggtg ttgatgacct
     2221 tcgtcgctta tgcatactca ggatgagttt tgtgaaaggc tggggaccgg attacccaag
     2281 acagagcatc aaagaaacac cttgctggat tgaaattcac ttacaccggg ccctccagct
     2341 cctagacgaa gtacttcata ccatgccgat tgcagaccca caacctttag actgaatcga
     2401 taaaataaaa gattttattt agtctccaga aaaagggggg aatgaaagac cccacctgta
     2461 ggtttggcaa gctagcttaa gtaacgccat tttgcaaggc atggaaaaat acataactga
     2521 gaatagagaa gttcagatca aggtcaggaa cagatggaac agctgaatat gggccaaaca
     2581 ggatatctgt ggtaagcagt tcctgccccg gctcagggcc aagaacagat ggaacagctg
     2641 aatatgggcc aaacaggata tctgtggtaa gcagttcctg ccccggctca gggccaagaa
     2701 cagatggtcc ccagatgcgg tccagccctc agcagtttct agagaaccat cagatgtttc
     2761 cagggtgccc caaggacctg aaatgaccct gtgccttatt tgaactaacc aatcagttcg
     2821 cttctcgctt ctgttcgcgc gcttctgctc cccgagctca ataaaagagc ccacaacccc
     2881 tcactcgggg cgccagtcct ccgattgact gagtcgcccg ggtacccgtg tatccaataa
     2941 accctcttgc agttgcatcc gacttgtggt ctcgctgttc cttgggaggg tctcctctga
     3001 gtgattgact acccgtcagc gggggtcttt catttggggg ctcgtccggg atcgggagac
     3061 ccctgcccag ggaccaccga cccaccaccg ggaggtaagc tggctgcctc gcgcgtttcg
     3121 gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt
     3181 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc
     3241 ggggcgcagc catgacccag tcacgtagcg atagcggagt gtatactggc ttaactatgc
     3301 ggcatcagag cagattgtac tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg
     3361 cgtaaggaga aaataccgca tcaggcgctc ttccgcttcc tcgctcactg actcgctgcg
     3421 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     3481 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     3541 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     3601 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     3661 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     3721 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     3781 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     3841 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     3901 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     3961 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt
     4021 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     4081 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattactcg
     4141 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     4201 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     4261 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     4321 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     4381 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     4441 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     4501 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     4561 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     4621 tttgcgcaac gttgttgcca ttgctgcagg catcgtggtg tcacgctcgt cgtttggtat
     4681 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     4741 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     4801 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     4861 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     4921 accgagttgc tcttgcccgg cgtcaacacg ggataatacc gcgccacata gcagaacttt
     4981 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     5041 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     5101 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     5161 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     5221 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     5281 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccattat
     5341 tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc ttcaagaatt
     5401 cataccagat caccgaaaac tgtcctccaa atgtgtcccc ctcacactcc caaattcgcg
     5461 ggcttctgcc tcttagacca ctctacccta ttccccacac tcaccggagc caaagccgcg
     5521 gcccttccgt ttctttgctt ttgaaagacc ccacccgtag gtggcaagct agcttaagta
     5581 acgccacttt gcaaggcatg gaaaaataca taactgagaa tagaaaagtt cagatcaagg
     5641 tcaggaacaa agaaacagct gaataccaaa caggatatct gtggtaagcg gttcctgccc
     5701 cggctcaggg ccaagaacag atgagacagc tgagtgatgg gccaaacagg atatctgtgg
     5761 taagcagttc ctgccccggc tcggggccaa gaacagatgg tccccagatg cggtccagcc
     5821 ctcagcagtt tctagtgaat catcagatgt ttccagggtg ccccaaggac ctgaaaatga
     5881 ccctgtacct tatttgaact aaccaatcag ttcgcttctc gcttctgttc gcgcgcttcc
     5941 gctctccgag ctcaataaaa gagcccacaa cccctcactc ggcgcgccag tcttccgata
     6001 gactgcgtcg cccgggtacc cgtattccca ataaagcctc ttgctgtttg catccgaatc
     6061 gtggtctcgc tgttccttgg gagggtctcc tctgagtgat tgactaccca cgacgggggt
     6121 ctttcatttg ggggctcgtc cgggatttgg agacccctgc ccagggacca ccgacccacc
     6181 accgggaggt aagctggcca gcaacttatc tgtgtctgtc cgattgtcta gtgtctatgt
     6241 ttgatgttat gcgcctgcgt ctgtactagt tagctaacta gctctgtatc tggcggaccc
     6301 gtggtggaac tgacgagttc tgaacacccg gccgcaaccc tgggagacgt cccagggact
     6361 ttgggggccg tttttgtggc ccgacctgag gaagggagtc gatgtggaat ccgaccccgt
     6421 caggatatgt ggttctggta ggagacgaga acctaaaaca gttcccgcct ccgtctgaat
     6481 ttttgctttc ggtttggaac cgaagccgcg cgtcttgtct gctgcagcgc tgcagcatcg
     6541 ttctgtgttg tctctgtctg actgtgtttc tgtatttgtc tgaaaattag ggccagactg
     6601 ttaccactcc cttaagtttg accttaggtc actggaaaga tgtcgagcgg atcgctcaca
     6661 accagtcggt agatgtcaag aagagacgtt gggttacctt ctgctctgca gaatggccaa
     6721 cctttaacgt cggatggccg cgagacggca cctttaaccg agacctcatc acccaggtta
     6781 agatcaaggt cttttcacct ggcccgcatg gacacccaga ccaggtcccc tacatcgtga
     6841 cctgggaagc cttggctttt gacccccctc cctgggtcaa gccctttgta caccctaagc
     6901 ctccgcctcc tcttcctcca tccgccccgt ctctccccct tgaacctcct cgttcgaccc
     6961 cgcctcgatc ctccctttat ccagccctca ctccttctct aggcgccgga attccgatct
     7021 gatcagctta ccatgaccga gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc
     7081 cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac gcgccacacc
     7141 gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt cctcacgcgc
     7201 gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg
     7261 accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc gcgcatggcc
     7321 gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct ggcgccgcac
     7381 cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga ccaccagggc
     7441 aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg cgccggggtg
     7501 cccgccttcc tggagacctc cgcgccccgc aacctcccct tctacgagcg gctcggcttc
     7561 accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag
     7621 cccggtgcct gacgcccgcc ccacgacccg cagcgcccga ccgaaaggag cgcacgaccc
     7681 catgcatcgg atccggccat tagccatatt attcattggt tatatagcat aaatcaatat
     7741 tggctattgg ccattgcata cgttgtatcc atatcataat atgtacattt atattggctc
     7801 atgtccaaca ttaccgccat gtt
//
