LOCUS       sequence-360240-        6417 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  N-terminally tagged Salmonella Typhimurium SopB (K528A) for 
            mammalian expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6417)
  AUTHORS   Walpole GFW, Pacheco J, Chauhan N, Clark J, Anderson KE, Abbas YM, 
            Brabant-Kirwan D, Montano-Rendon F, Liu Z, Zhu H, Brumell JH, 
            Deiters A, Stephens LR, Hawkins PT, Hammond GRV, Grinstein S, Fairn 
            GD
  TITLE     Kinase-independent synthesis of 3-phosphorylated phosphoinositides 
            by a phosphotransferase.
  JOURNAL   Nat Cell Biol. 2022 Apr 28. pii: 10.1038/s41556-022-00895-y. doi: 
            10.1038/s41556-022-00895-y.
  PUBMED    35484249
REFERENCE   2  (bases 1 to 6417)
  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"; journalName: "Nat Cell 
            Biol. 2022 Apr 28. pii: 10.1038/s41556-022-00895-y. doi: 
            10.1038/s41556-022-00895-y."
COMMENT     Sequence Label: EGFP-SopB(K528A)
FEATURES             Location/Qualifiers
     source          1..6417
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        74..377
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        378..581
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     532..552
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      620..629
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(626..628,629..631,632..1342)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(626..628,629..631,632..1342)
                     /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(671..692)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(932..951)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1279..1300
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3218..3339
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3255..3274)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3309..3328
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3346..3801)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3483..3504)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3695..3714
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3828..3932
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3934..4291
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3934..4125
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3957..3977)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4142..4277
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4326..5120
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4326..5120
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4380..4399)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4990..5009
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5308..5327)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5352..5399
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5728..6316
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6217..6236
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 taccgccatg cattagttat taatagtaat caattacggg gtcattagtt catagcccat
       61 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      121 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      181 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      241 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      301 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      361 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      421 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      481 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      541 gcggtaggcg tgtacggtgg gaggtctata taagcagagc tggtttagtg aaccgtcaga
      601 tccgctagcg ctaccggtcg ccaccatggt gagcaagggc gaggagctgt tcaccggggt
      661 ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg
      721 cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg
      781 caagctgccc gtgccctggc ccaccctcgt gaccaccctg acctacggcg tgcagtgctt
      841 cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg
      901 ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga
      961 ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa
     1021 ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta
     1081 tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat
     1141 cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg
     1201 ccccgtgctg ctgcccgaca accactacct gagcacccag tccgccctga gcaaagaccc
     1261 caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct
     1321 cggcatggac gagctgtaca agtccggact cagatctcga gctcaagctt cgaattctgc
     1381 agtcgacggt ggtagtgctt ccggacaaat acagagcttc tatcactcag cttcactaaa
     1441 aacccaggag gcttttaaaa gcctacaaaa aaccttatac aacggaatgc agattctctc
     1501 aggccagggc aaagcgccgg ctaaagcgcc cgacgctcgc ccggaaatta ttgtcctgcg
     1561 agaacccggc gcgacatggg ggaattatct acagcatcag aaggcgtcta accactcgct
     1621 gcataacctc tataacttac agcgcgatct tcttaccgtc gcggcaaccg ttctgggtaa
     1681 acaagacccg gttctaacgt caatggcaaa ccaaatggag ttagccaaag ttaaagcgga
     1741 ccggccagca acaaaacaag aagaagccgc ggcaaaagca ttgaagaaaa atcttatcga
     1801 acttattgca gcacgcactc agcagcagga tggcttacct gcaaaagaag ctcatcgctt
     1861 tgcggcagta gcgtttagag atgctcaggt caagcagctt aataaccagc cctggcaaac
     1921 cataaaaaat acactcacgc ataacgggca tcactatacc aacacgcagc tccctgcagc
     1981 agagatgaaa atcggcgcaa aagatatctt tcccagtgct tatgagggaa agggcgtatg
     2041 cagttgggat accaagaata ttcatcacgc caataatttg tggatgtcca cggtgagtgt
     2101 gcatgaggac ggtaaagata aaacgctttt ttgcgggata cgtcatggcg tgctttcccc
     2161 ctatcatgaa aaagatccgc ttctgcgtca cgtcggcgct gaaaacaaag ccaaagaagt
     2221 attaactgcg gcacttttta gtaaacctga gttgcttaac aaagccttag cgggcgaggc
     2281 ggtaagcctg aaactggtat ccgtcgggtt actcaccgcg tcgaatattt tcggcaaaga
     2341 gggaacgatg gtcgaggacc aaatgcgcgc atggcaatcg ttgacccagc cgggaaaaat
     2401 gattcattta aaaatccgca ataaagatgg cgatctacag acggtaaaaa taaaaccgga
     2461 cgtcgccgca tttaatgtgg gtgttaatga gctggcgctc aagctcggct ttggccttaa
     2521 ggcatcggat agctataatg ccgaggcgct acatcagtta ttaggcaatg atttacgccc
     2581 tgaagccaga ccaggtggct gggttggcga atggctggcg caatacccgg ataattatga
     2641 ggtcgtcaat acattagcgc gccagattaa ggatatatgg aaaaataacc aacatcataa
     2701 agatggcggc gaaccctata aactcgcaca acgccttgcc atgttagccc atgaaattga
     2761 cgcggtaccc gcctggaatt gtaaaagcgg caaagatcgt acagggatga tggattcaga
     2821 aatcaagcga gagatcattt ccttacatca gacccatatg ttaagtgcgc ctggtagtct
     2881 tccggatagc ggtggacaga aaattttcca aaaagtatta ctgaatagcg gtaacctgga
     2941 gattcagaaa caaaatacgg gcggggcggg aaacaaagta atggcgaatt tatcgccaga
     3001 ggtgctcaat ctttcctatc aaaaacgagt tggggatgaa aatatttggc agtcagtaaa
     3061 aggcatttct tcattaatca catcttaagg atccaccgga tctagataac tgatcataat
     3121 cagccatacc acatttgtag aggttttact tgctttaaaa aacctcccac acctccccct
     3181 gaacctgaaa cataaaatga atgcaattgt tgttgttaac ttgtttattg cagcttataa
     3241 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     3301 ttctagttgt ggtttgtcca aactcatcaa tgtatcttaa cgcgtaaatt gtaagcgtta
     3361 atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg
     3421 ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg
     3481 ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa
     3541 aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg
     3601 ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt
     3661 gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg
     3721 ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta
     3781 atgcgccgct acagggcgcg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     3841 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     3901 aaatgcttca ataatattga aaaaggaaga gtcctgaggc ggaaagaacc agctgtggaa
     3961 tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     4021 catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag
     4081 aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc
     4141 catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     4201 ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg
     4261 aggctttttt ggaggcctag gcttttgcaa agatcgatca agagacagga tgaggatcgt
     4321 ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc
     4381 tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc
     4441 tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg
     4501 aactgcaaga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag
     4561 ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg
     4621 ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg
     4681 caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac
     4741 atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg
     4801 acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgagcatgc
     4861 ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg
     4921 aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc
     4981 aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc
     5041 gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc
     5101 ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc
     5161 caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg
     5221 aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt
     5281 cttcgcccac cctaggggga ggctaactga aacacggaag gagacaatac cggaaggaac
     5341 ccgcgctatg acggcaataa aaagacagaa taaaacgcac ggtgttgggt cgtttgttca
     5401 taaacgcggg gttcggtccc agggctggca ctctgtcgat accccaccga gaccccattg
     5461 gggccaatac gcccgcgttt cttccttttc cccaccccac cccccaagtt cgggtgaagg
     5521 cccagggctc gcagccaacg tcggggcggc aggccctgcc atagcctcag gttactcata
     5581 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     5641 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     5701 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     5761 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     5821 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     5881 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     5941 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     6001 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     6061 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     6121 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     6181 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     6241 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     6301 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     6361 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtat
//
