LOCUS       Exported                8456 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 8456)
  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 8456)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8456
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     LTR             414..589
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    652..851
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1052..1468
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1398..1420
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1436..1452
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     CDS             1512..2306
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1566..1585)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2176..2195
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     enhancer        2377..2680
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2681..2884
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2835..2855
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             2931..3647
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             2931..3647
                     /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(2976..2997)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3237..3256)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3584..3605
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             5358..5951
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(6054..6076)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    6162..6302
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     6217..6236
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(6317..6334)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6488..7076)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6568..6587)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(7247..8107)
                     /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     7870..7889
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8108..8212)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     8280..8298
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tttgaaagac cccacccgta ggtggcaagc tagcttaagt aacgccactt tgcaaggcat
       61 ggaaaaatac ataactgaga atagaaaagt tcagatcaag gtcaggaaca aagaaacagc
      121 tgaataccaa acaggatatc tgtggtaagc ggttcctgcc ccggctcagg gccaagaaca
      181 gatgagacag ctgagtgatg ggccaaacag gatatctgtg gtaagcagtt cctgccccgg
      241 ctcggggcca agaacagatg gtccccagat gcggtccagc cctcagcagt ttctagtgaa
      301 tcatcagatg tttccagggt gccccaagga cctgaaaatg accctgtacc ttatttgaac
      361 taaccaatca gttcgcttct cgcttctgtt cgcgcgcttc cgctctccga gctcaataaa
      421 agagcccaca acccctcact cggcgcgcca gtcttccgat agactgcgtc gcccgggtac
      481 ccgtattccc aataaagcct cttgctgttt gcatccgaat cgtggtctcg ctgttccttg
      541 ggagggtctc ctctgagtga ttgactaccc acgacggggg tctttcattt gggggctcgt
      601 ccgggatttg gagacccctg cccagggacc accgacccac caccgggagg taagctggcc
      661 agcaacttat ctgtgtctgt ccgattgtct agtgtctatg tttgatgtta tgcgcctgcg
      721 tctgtactag ttagctaact agctctgtat ctggcggacc cgtggtggaa ctgacgagtt
      781 ctgaacaccc ggccgcaacc ctgggagacg tcccagggac tttgggggcc gtttttgtgg
      841 cccgacctga ggaagggagt cgatgtggaa tccgaccccg tcaggatatg tggttctggt
      901 aggagacgag aacctaaaac agttcccgcc tccgtctgaa tttttgcttt cggtttggaa
      961 ccgaagccgc gcgtcttgtc tgctgcagcg ctgcagcatc gttctgtgtt gtctctgtct
     1021 gactgtgttt ctgtatttgt ctgaaaatta gggccagact gttaccactc ccttaagttt
     1081 gaccttaggt cactggaaag atgtcgagcg gatcgctcac aaccagtcgg tagatgtcaa
     1141 gaagagacgt tgggttacct tctgctctgc agaatggcca acctttaacg tcggatggcc
     1201 gcgagacggc acctttaacc gagacctcat cacccaggtt aagatcaagg tcttttcacc
     1261 tggcccgcat ggacacccag accaggtccc ctacatcgtg acctgggaag ccttggcttt
     1321 tgacccccct ccctgggtca agccctttgt acaccctaag cctccgcctc ctcttcctcc
     1381 atccgccccg tctctccccc ttgaacctcc tcgttcgacc ccgcctcgat cctcccttta
     1441 tccagccctc actccttctc taggcgccgg aattccgatc tgatcaagag acaggatgag
     1501 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     1561 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     1621 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     1681 tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
     1741 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     1801 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     1861 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     1921 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     1981 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc
     2041 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     2101 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     2161 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
     2221 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     2281 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaatagt aatcaattac
     2341 ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg
     2401 cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc
     2461 catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac
     2521 tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa
     2581 tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac
     2641 ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta
     2701 catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga
     2761 cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa
     2821 ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag
     2881 agctggttta gtgaaccgtc agatccgcta gcgctaccgg actcagatcc atggtgagca
     2941 agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa
     3001 acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga
     3061 ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca
     3121 ccctgaccta cggcgtgcag tgcttcagcc gctaccccga ccacatgaag cagcacgact
     3181 tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg
     3241 acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca
     3301 tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt
     3361 acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac ggcatcaagg
     3421 tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc gaccactacc
     3481 agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagca
     3541 cccagtccgc cctgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt
     3601 tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagaga tccatggaca
     3661 atatgtctat tacgaataca ccaacaagta atgatgcctg tctgagcatt gtgcatagtt
     3721 tgatgtgcca tagacaaggt ggagagagtg aaacatttgc aaaaagagca attgaaagtt
     3781 tggtaaagaa gctgaaggag aaaaaagatg aattggattc tttaataaca gctataacta
     3841 caaatggagc tcatcctagt aaatgtgtta ccatacagag aacattggat gggaggcttc
     3901 aggtggctgg tcggaaagga tttcctcatg tgatctatgc ccgtctctgg aggtggcctg
     3961 atcttcacaa aaatgaacta aaacatgtta aatattgtca gtatgcgttt gacttaaaat
     4021 gtgatagtgt ctgtgtgaat ccatatcact acgaacgagt tgtatcacct ggaattgatc
     4081 tctcaggatt aacactgcag agtaatgctc catcaagtat gatggtgaag gatgaatacg
     4141 ttcatgactt tgagggacag ccatcgttgt ccactgaagg acattcaatt caaaccatcc
     4201 agcatccacc aagtaatcgt gcatcgacag agacatacag caccccagct ctgttagccc
     4261 catctgagtc taatgctacc agcactgcca actttcccaa cattcctgtg gcttccacaa
     4321 gtcagcctgc cagtatactg gggggcagcc atagtgaagg actgttgcag atagcatcag
     4381 ggcctcagcc aggacagcag cagaatggat ttactggtca gccagctact taccatcata
     4441 acagcactac cacctggact ggaagtagga ctgcaccata cacacctaat ttgcctcacc
     4501 accaaaacgg ccatcttcag caccacccgc ctatgccgcc ccatcccgga cattactggc
     4561 ctgttcacaa tgagcttgca ttccagcctc ccatttccaa tcatcctgct cctgagtatt
     4621 ggtgttccat tgcttacttt gaaatggatg ttcaggtagg agagacattt aaggttcctt
     4681 caagctgccc tattgttact gttgatggat acgtggaccc ttctggagga gatcgctttt
     4741 gtttgggtca actctccaat gtccacagga cagaagccat tgagagagca aggttgcaca
     4801 taggcaaagg tgtgcagttg gaatgtaaag gtgaaggtga tgtttgggtc aggtgcctta
     4861 gtgaccacgc ggtctttgta cagagttact acttagacag agaagctggg cgtgcacctg
     4921 gagatgctgt tcataagatc tacccaagtg catatataaa ggtctttgat ttgcgtcagt
     4981 gtcatcgaca gatgcagcag caggcggcta ctgcacaagc tgcagcagct gcccaggcag
     5041 cagccgtggc aggaaacatc cctggcccag gatcagtagg tggaatagct ccagctatca
     5101 gtctgtcagc tgctgctgga attggtgttg atgaccttcg tcgcttatgc atactcagga
     5161 tgagttttgt gaaaggctgg ggaccggatt acccaagaca gagcatcaaa gaaacacctt
     5221 gctggattga aattcactta caccgggccc tccagctcct agacgaagta cttcatacca
     5281 tgccgattgc agacccacaa cctttagact gaatcgataa aataaaagat tttatttagt
     5341 ctccagaaaa aggggggaat gaaagacccc acctgtaggt ttggcaagct agcttaagta
     5401 acgccatttt gcaaggcatg gaaaaataca taactgagaa tagagaagtt cagatcaagg
     5461 tcaggaacag atggaacagc tgaatatggg ccaaacagga tatctgtggt aagcagttcc
     5521 tgccccggct cagggccaag aacagatgga acagctgaat atgggccaaa caggatatct
     5581 gtggtaagca gttcctgccc cggctcaggg ccaagaacag atggtcccca gatgcggtcc
     5641 agccctcagc agtttctaga gaaccatcag atgtttccag ggtgccccaa ggacctgaaa
     5701 tgaccctgtg ccttatttga actaaccaat cagttcgctt ctcgcttctg ttcgcgcgct
     5761 tctgctcccc gagctcaata aaagagccca caacccctca ctcggggcgc cagtcctccg
     5821 attgactgag tcgcccgggt acccgtgtat ccaataaacc ctcttgcagt tgcatccgac
     5881 ttgtggtctc gctgttcctt gggagggtct cctctgagtg attgactacc cgtcagcggg
     5941 ggtctttcat ttgggggctc gtccgggatc gggagacccc tgcccaggga ccaccgaccc
     6001 accaccggga ggtaagctgg ctgcctcgcg cgtttcggtg atgacggtga aaacctctga
     6061 cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg gagcagacaa
     6121 gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gcgcagccat gacccagtca
     6181 cgtagcgata gcggagtgta tactggctta actatgcggc atcagagcag attgtactga
     6241 gagtgcacca tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca
     6301 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     6361 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     6421 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     6481 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     6541 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     6601 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     6661 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     6721 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     6781 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     6841 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     6901 ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc
     6961 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     7021 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     7081 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     7141 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     7201 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     7261 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     7321 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     7381 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     7441 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     7501 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     7561 ctgcaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     7621 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     7681 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     7741 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     7801 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     7861 caacacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     7921 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     7981 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     8041 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     8101 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     8161 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     8221 cccgaaaagt gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa
     8281 ataggcgtat cacgaggccc tttcgtcttc aagaattcat accagatcac cgaaaactgt
     8341 cctccaaatg tgtccccctc acactcccaa attcgcgggc ttctgcctct tagaccactc
     8401 taccctattc cccacactca ccggagccaa agccgcggcc cttccgtttc tttgct
//
