LOCUS       Exported                7894 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Plasmid encodes Strep-tagged full-length (unprocessed) VEGF-D of 
            human origin to be used for stimulation of lymphangiogenesis in mice
            in vivo..
ACCESSION   .
VERSION     .
KEYWORDS    psubCAG-WPRE-hVEGF-D-Strept
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7894)
  TITLE     Alitalo expression plasmids
REFERENCE   2  (bases 1 to 7894)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7894
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   35..164
                     /label=AAV2 ITR
     enhancer        223..602
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        604..881
                     /label=chicken beta-actin promoter
     intron          882..1898
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1822..1844
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1906..1925
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     intron          1989..2085
                     /label=SV40 intron
                     /note="modified SV40 intron with splice donor and acceptor 
                     sites"
     CDS             3218..3307
                     /codon_start=1
                     /product="two Strep-Tag(R)II moieties connected by a linker
                     for enhanced binding to Strep-Tactin(R), an engineered form
                     of streptavidin (Schmidt et al., 2013)"
                     /label=Twin-Strep-tag
                     /translation="SAWSHPQFEKGGGSGGGSGGSAWSHPQFEK"
     misc_feature    3321..3909
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3374..3394)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3792..3803)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3924..4058
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3948..3967)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4002..4021
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4208..4337
                     /label=AAV2 ITR
     rep_origin      4708..5221
                     /direction=RIGHT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4795..4814)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5005..5026
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5703..5722)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5822..5844
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5882..5900)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5968..6072
                     /gene="bla"
                     /label=AmpR promoter
     CDS             6073..6933
                     /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     complement(6291..6310)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7104..7692
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7593..7612
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7846..7863
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 ctctccccgc gcgttggccg attcattaat gcagctgcgc gctcgctcgc tcactgaggc
       61 cgcccgggca aagcccgggc gtcgggcgac ctttggtcgc ccggcctcag tgagcgagcg
      121 agcgcgcaga gagggagtgg ccaactccat cactaggggt tccttgtagt taatgattaa
      181 cccgccatgc tacttatcta cgtagccatg ctctagaatt tcgacattga ttattgacta
      241 gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg
      301 ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga
      361 cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat
      421 gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa
      481 gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca
      541 tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc gctattacca
      601 tggtcgaggt gagccccacg ttctgcttca ctctccccat ctcccccccc tccccacccc
      661 caattttgta tttatttatt ttttaattat tttgtgcagc gatgggggcg gggggggggg
      721 gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg gggcggggcg aggcggagag
      781 gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt tccttttatg gcgaggcggc
      841 ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc gggagtcgct gcgcgctgcc
      901 ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc gccccggctc tgactgaccg
      961 cgttactccc acaggtgagc gggcgggacg gcccttctcc tccgggctgt aattagcgct
     1021 tggtttaatg acggcttgtt tcttttctgt ggctgcgtga aagccttgag gggctccggg
     1081 agggcccttt gtgcgggggg agcggctcgg ggggtgcgtg cgtgtgtgtg tgcgtgggga
     1141 gcgccgcgtg cggctccgcg ctgcccggcg gctgtgagcg ctgcgggcgc ggcgcggggc
     1201 tttgtgcgct ccgcagtgtg cgcgagggga gcgcggccgg gggcggtgcc ccgcggtgcg
     1261 gggggggctg cgaggggaac aaaggctgcg tgcggggtgt gtgcgtgggg gggtgagcag
     1321 ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca cccccctccc cgagttgctg
     1381 agcacggccc ggcttcgggt gcggggctcc gtacggggcg tggcgcgggg ctcgccgtgc
     1441 cgggcggggg gtggcggcag gtgggggtgc cgggcggggc ggggccgcct cgggccgggg
     1501 agggctcggg ggaggggcgc ggcggccccc ggagcgccgg cggctgtcga ggcgcggcga
     1561 gccgcagcca ttgcctttta tggtaatcgt gcgagagggc gcagggactt cctttgtccc
     1621 aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac cccctctagc gggcgcgggg
     1681 cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga gggccttcgt gcgtcgccgc
     1741 gccgccgtcc ccttctccct ctccagcctc ggggctgtcc gcggggggac ggctgccttc
     1801 gggggggacg gggcagggcg gggttcggct tctggcgtgt gaccggcggc tctagagcct
     1861 ctgctaacca tgttcatgcc ttcttctttt tcctacagct cctgggcaac gtgctggtta
     1921 ttgtgctgtc tcatcatttt ggcaaagaat tcctcgatcg aggaactgaa aaaccagaaa
     1981 gttaactggt aagtttagtc tttttgtctt ttatttcagg tcccggatcc ggtggtggtg
     2041 caaatcaaag aactgctcct cagtggatgt tgcctttact tctaggcctg tacggaagtg
     2101 ttacttctgc tctaaaagct gcggaattgt acccgcggcc gctagcgcca ccatgtacag
     2161 agagtgggta gtggtgaatg ttttcatgat gttgtacgtc cagctggtgc agggctccag
     2221 taatgaacat ggaccagtga agcgatcatc tcagtccaca ttggaacgat ctgaacagca
     2281 gatcagggct gcttctagtt tggaggaact acttcgaatt actcactctg aggactggaa
     2341 gctgtggaga tgcaggctga ggctcaaaag ttttaccagt atggactctc gctcagcatc
     2401 ccatcggtcc actaggtttg cggcaacttt ctatgacatt gaaacactaa aagttataga
     2461 tgaagaatgg caaagaactc agtgcagccc tagagaaacg tgcgtggagg tggccagtga
     2521 gctggggaag agtaccaaca cattcttcaa gcccccttgt gtgaacgtgt tccgatgtgg
     2581 tggctgttgc aatgaagaga gccttatctg tatgaacacc agcacctcgt acatttccaa
     2641 acagctcttt gagatatcag tgcctttgac atcagtacct gaattagtgc ctgttaaagt
     2701 tgccaatcat acaggttgta agtgcttgcc aacagccccc cgccatccat actcaattat
     2761 cagaagatcc atccagatcc ctgaagaaga tcgctgttcc cattccaaga aactctgtcc
     2821 tattgacatg ctatgggata gcaacaaatg taaatgtgtt ttgcaggagg aaaatccact
     2881 tgctggaaca gaagaccact ctcatctcca ggaaccagct ctctgtgggc cacacatgat
     2941 gtttgacgaa gatcgttgcg agtgtgtctg taaaacacca tgtcccaaag atctaatcca
     3001 gcaccccaaa aactgcagtt gctttgagtg caaagaaagt ctggagacct gctgccagaa
     3061 gcacaagcta tttcacccag acacctgcag ctgtgaggac agatgcccct ttcataccag
     3121 accatgtgca agtggcaaaa cagcatgtgc aaagcattgc cgctttccaa aggagaaaag
     3181 ggctgcccag gggccccaca gccgaaagaa tcctgggagc gcttggagcc acccacagtt
     3241 cgaaaaaggt ggaggttctg gcggtggatc gggaggttca gcgtggtctc atcctcaatt
     3301 tgagaagtga acgcgtcgat aatcaacctc tggattacaa aatttgtgaa agattgactg
     3361 gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt
     3421 atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc
     3481 tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt
     3541 ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga
     3601 ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct
     3661 gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaagctga
     3721 cgtcctttcc atggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct
     3781 gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc
     3841 tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg
     3901 cctccccgca tcgcggccgc ggggatccag acatgataag atacattgat gagtttggac
     3961 aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg
     4021 ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt
     4081 ttatgtttca ggttcagggg gaggtgtggg aggttttttc ggatcctcta gagtcgacct
     4141 gcaggcatgc aagctctaga gcatggctac gtagataagt agcatggcgg gttaatcatt
     4201 aactacaagg aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc
     4261 actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg
     4321 agcgagcgag cgcgcagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa
     4381 cagttgcgca gcctgaatgg cgaatggcga ttccgttgca atggctggcg gtaatattgt
     4441 tctggatatt accagcaagg ccgatagttt gagttcttct actcaggcaa gtgatgttat
     4501 tactaatcaa agaagtattg cgacaacggt taatttgcgt gatggacaga ctcttttact
     4561 cggtggcctc actgattata aaaacacttc tcaggattct ggcgtaccgt tcctgtctaa
     4621 aatcccttta atcggcctcc tgtttagctc ccgctctgat tctaacgagg aaagcacgtt
     4681 atacgtgctc gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta agcgcggcgg
     4741 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     4801 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     4861 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     4921 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     4981 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     5041 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     5101 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa
     5161 tttaaatatt tgcttataca atcttcctgt ttttggggct tttctgatta tcaaccgggg
     5221 tacatatgat tgacatgcta gttttacgat taccgttcat cgattctctt gtttgctcca
     5281 gactctcagg caatgacctg atagcctttg tagagacctc tcaaaaatag ctaccctctc
     5341 cggcatgaat ttatcagcta gaacggttga atatcatatt gatggtgatt tgactgtctc
     5401 cggcctttct cacccgtttg aatctttacc tacacattac tcaggcattg catttaaaat
     5461 atatgagggt tctaaaaatt tttatccttg cgttgaaata aaggcttctc ccgcaaaagt
     5521 attacagggt cataatgttt ttggtacaac cgatttagct ttatgctctg aggctttatt
     5581 gcttaatttt gctaattctt tgccttgcct gtatgattta ttggatgttg gaatcgcctg
     5641 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatatg gtgcactctc
     5701 agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct
     5761 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
     5821 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag
     5881 ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg
     5941 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     6001 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     6061 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     6121 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     6181 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     6241 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     6301 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     6361 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     6421 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     6481 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     6541 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     6601 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     6661 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     6721 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     6781 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     6841 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     6901 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     6961 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     7021 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     7081 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     7141 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     7201 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     7261 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     7321 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     7381 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     7441 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     7501 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     7561 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     7621 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     7681 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     7741 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     7801 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     7861 gaggaagcgg aagagcgccc aatacgcaaa ccgc
//
