LOCUS       sequence-389343-        7786 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression vector containing GFP tagged Podocalyxin. PKC 
            phosphorylation-deficient mutant..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7786)
  AUTHORS   Bryant DM, Roignot J, Datta A, Overeem AW, Kim M, Yu W, Peng X, 
            Eastburn DJ, Ewald AJ, Werb Z, Mostov KE
  TITLE     A molecular switch for the orientation of epithelial cell 
            polarization.
  JOURNAL   Dev Cell. 2014 Oct 27;31(2):171-87. doi: 
            10.1016/j.devcel.2014.08.027. Epub 2014 Oct 9.
  PUBMED    25307480
REFERENCE   2  (bases 1 to 7786)
  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.devcel.2014.08.027"; journalName: "Dev Cell"; date: 
            "2014-10-27- 27"; volume: "31"; issue: "2"; pages: "171-87"
COMMENT     Sequence Label: GFP-PODXL S481A
FEATURES             Location/Qualifiers
     source          1..7786
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(62..81)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        253..632
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        633..836
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     787..807
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     881..900
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        881..899
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             982..1014
                     /codon_start=1
                     /product="epitope tag from vesicular stomatitis virus G 
                     protein"
                     /label=VSV-G tag
                     /translation="YTDIEMNRLGK"
     CDS             join(1015..1017,1018..1020,1021..1731)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMTDKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1060..1081)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1321..1340)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1668..1689
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(3353..3371)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3354..3371)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3391..3408)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3397..3621
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3667..4095
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3754..3773)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3964..3985
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4104..4124)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4109..4438
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4289..4424
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4351..4370
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4505..5299
                     /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(4559..4578)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5169..5188
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5473..5594
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5510..5529)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5564..5583
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5643..5659)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5643..5659)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5656..5678)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5667..5683
                     /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(5691..5697,5698..5715,5716..5721))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5736..5757
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5874..5891)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6045..6633)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6125..6144)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6804..7595,7596..7664))
                     /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     7427..7446
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7665..7769)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tgccacctga cgtcgacgga tcgggagatc gatctcccga tcccctatgg tgcactctca
       61 gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct tgtgtgttgg
      121 aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc ttgaccgaca
      181 attgcatgaa gaatctgctt agggttaggc gttttgcgct gcttcgcgat gtacgggcca
      241 gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat
      301 tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg
      361 gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa
      421 cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact
      481 tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta
      541 aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt
      601 acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg
      661 ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg
      721 ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc
      781 cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctctct
      841 ggctaactag agaacccact gcttactggc ttatcgaaat taatacgact cactataggg
      901 agacccaagc ttggtaccat ggccttacca gtgaccgcct tgctcctgcc gctagccttg
      961 ctgctgcatg ccgccaggcc gtatactgat atcgaaatga atcgattggg taagatggtg
     1021 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac
     1081 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag
     1141 ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg
     1201 accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac
     1261 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag
     1321 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
     1381 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
     1441 gagtacaact acaacagcca caacgtctat atcatgaccg acaagcagaa gaacggcatc
     1501 aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac
     1561 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     1621 agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     1681 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa gaatggcaca
     1741 gaaggcccta acttctacgt gcccttctcc aatgcgacgg aattccagga aaagagtccc
     1801 cagcccggcc caacccccat ggccacttca acatctacca ggcctgcccc agcctcagcc
     1861 ccagccccca aaagctccgt ggcggcttcg gtcccggccg agcagaacac tacacccatg
     1921 acgacgaaag ccccagccac ccagagcccc tctgcatccc ctggaagctc cgtggagaac
     1981 tcagccccgg cccagggctc gaccacaaca cagcagagcc tgagtgttac tactaaggct
     2041 gaagctaagg atgcaggtgg cgtccccacg gcacacgtca ccggctctgc tcggcctgtc
     2101 accagtggca gccaggtcgc agctcaggac ccagctgcat ccaaagctcc gagcaaccac
     2161 agcatcacca caaagcctct ggccacagag gccacgagcc aggccccccg tcagaccaca
     2221 gatgtcggca ccccgggacc cactgctcct cctgtgacca actcgaccag ccctgacctg
     2281 ctgggccatg ccaccccgaa gccctccgag gggccccagc tcagtttccc cacggctgct
     2341 ggcagcctgg gccctgtcac tggctctggc accggctctg gcaccctctc cacaccccag
     2401 gggaagcctg cgacactgac gcccgtggcc agctcagcag agactcaagg gatgcccagc
     2461 ccgatgccac ccagcccagc ctccccctcc tcctccccct ttccctcctc tccctccccc
     2521 tcccccgcac tgcagccctc gggcccctcg gcggcaggaa cagaggacac aaccggcagg
     2581 ggccccacaa gctccagcac tgagttggct tcaactgccc tgcacggccc cagcacgctt
     2641 tcccccacct cggcagtgag agaccagagg gtaagctgcg ggcctccgga gagaccaacc
     2701 gagcagctgc tcatcctgaa cctcaccaga agcagcccct gcacagacag ccttccggaa
     2761 gacaaactgg tcaccctgct gtgccgagcg gccaagccca ccttcaaccc cgcccaggac
     2821 cagtgccacg tgctgctggc gcccatgctg gggagccacg cggtggtggt caaagaaatc
     2881 accatcaaga cgaatctgct gcccacggcc gtgttcgagc tgctgaagga cagatgggat
     2941 gacctgagag aggagggcgt cagcgacatg cagctggggg accaggggcc cccggaggag
     3001 accgaggacc gcttcagcct gccgctcatc atcaccatcg tctgcatggc ctccttcctg
     3061 ctgcttgtcg cggccctcta cggctgctgc caccagcgcc tcgcccatag gaaggaccag
     3121 caacgactca cggaggagct gcagaccgtg gagaacggtt accatgacaa cccgacccta
     3181 gaagtgatgg agacgtcggc cgagatgcag gagaagaagg tggtcaacct taacggggag
     3241 ctgggggaca gctggattgt cccgctggac aacctgacca aggacgacct ggacgaggag
     3301 gaggacacgc acctctaagc ggccgctcga gcatgcatct agagggccct attctatagt
     3361 gtcacctaaa tgctagagct cgctgatcag cctcgactgt gccttctagt tgccagccat
     3421 ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc
     3481 tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg
     3541 ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg
     3601 gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt
     3661 atccccacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     3721 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     3781 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     3841 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     3901 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     3961 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     4021 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     4081 aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     4141 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     4201 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     4261 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     4321 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc
     4381 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     4441 gctcccggga gcttgtatat ccattttcgg atctgatcaa gagacaggat gaggatcgtt
     4501 tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct
     4561 attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct
     4621 gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga
     4681 actgcaggac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc
     4741 tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg
     4801 gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc
     4861 aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca
     4921 tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga
     4981 cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg cgcgcatgcc
     5041 cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga
     5101 aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca
     5161 ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg
     5221 cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct
     5281 tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc
     5341 aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga
     5401 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc
     5461 ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     5521 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     5581 atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca
     5641 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     5701 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     5761 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     5821 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     5881 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     5941 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     6001 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     6061 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     6121 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     6181 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     6241 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     6301 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     6361 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     6421 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     6481 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     6541 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     6601 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     6661 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     6721 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     6781 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     6841 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     6901 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     6961 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     7021 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     7081 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     7141 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     7201 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     7261 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     7321 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     7381 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     7441 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     7501 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     7561 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     7621 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     7681 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     7741 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaag
//
