LOCUS       sequence-360236-        6417 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  N-terminally tagged Salmonella Typhimurium SopB (K525A) 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(K525A)
FEATURES             Location/Qualifiers
     source          1..6417
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        76..379
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        380..583
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     534..554
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      622..631
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(628..630,631..633,634..1344)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(628..630,631..633,634..1344)
                     /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(673..694)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(934..953)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1281..1302
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3220..3341
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3257..3276)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3311..3330
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3348..3803)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3485..3506)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3697..3716
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3830..3934
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3936..4293
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3936..4127
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3959..3979)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4144..4279
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4328..5122
                     /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             4328..5122
                     /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(4382..4401)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4992..5011
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5310..5329)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5354..5401
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5730..6318
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6219..6238
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 attaccgcca tgcattagtt attaatagta atcaattacg gggtcattag ttcatagccc
       61 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
      121 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
      181 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
      241 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
      301 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
      361 agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg
      421 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
      481 gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat
      541 gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctggtttag tgaaccgtca
      601 gatccgctag cgctaccggt cgccaccatg gtgagcaagg gcgaggagct gttcaccggg
      661 gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc
      721 ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc
      781 ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc
      841 ttcagccgct accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa
      901 ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc
      961 gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc
     1021 aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc
     1081 tatatcatgg ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac
     1141 atcgaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac
     1201 ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccgccct gagcaaagac
     1261 cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact
     1321 ctcggcatgg acgagctgta caagtccgga ctcagatctc gagctcaagc ttcgaattct
     1381 gcagtcgacg gtggtagtgc ttccggacaa atacagagct tctatcactc agcttcacta
     1441 aaaacccagg aggcttttaa aagcctacaa aaaaccttat acaacggaat gcagattctc
     1501 tcaggccagg gcaaagcgcc ggctaaagcg cccgacgctc gcccggaaat tattgtcctg
     1561 cgagaacccg gcgcgacatg ggggaattat ctacagcatc agaaggcgtc taaccactcg
     1621 ctgcataacc tctataactt acagcgcgat cttcttaccg tcgcggcaac cgttctgggt
     1681 aaacaagacc cggttctaac gtcaatggca aaccaaatgg agttagccaa agttaaagcg
     1741 gaccggccag caacaaaaca agaagaagcc gcggcaaaag cattgaagaa aaatcttatc
     1801 gaacttattg cagcacgcac tcagcagcag gatggcttac ctgcaaaaga agctcatcgc
     1861 tttgcggcag tagcgtttag agatgctcag gtcaagcagc ttaataacca gccctggcaa
     1921 accataaaaa atacactcac gcataacggg catcactata ccaacacgca gctccctgca
     1981 gcagagatga aaatcggcgc aaaagatatc tttcccagtg cttatgaggg aaagggcgta
     2041 tgcagttggg ataccaagaa tattcatcac gccaataatt tgtggatgtc cacggtgagt
     2101 gtgcatgagg acggtaaaga taaaacgctt ttttgcggga tacgtcatgg cgtgctttcc
     2161 ccctatcatg aaaaagatcc gcttctgcgt cacgtcggcg ctgaaaacaa agccaaagaa
     2221 gtattaactg cggcactttt tagtaaacct gagttgctta acaaagcctt agcgggcgag
     2281 gcggtaagcc tgaaactggt atccgtcggg ttactcaccg cgtcgaatat tttcggcaaa
     2341 gagggaacga tggtcgagga ccaaatgcgc gcatggcaat cgttgaccca gccgggaaaa
     2401 atgattcatt taaaaatccg caataaagat ggcgatctac agacggtaaa aataaaaccg
     2461 gacgtcgccg catttaatgt gggtgttaat gagctggcgc tcaagctcgg ctttggcctt
     2521 aaggcatcgg atagctataa tgccgaggcg ctacatcagt tattaggcaa tgatttacgc
     2581 cctgaagcca gaccaggtgg ctgggttggc gaatggctgg cgcaataccc ggataattat
     2641 gaggtcgtca atacattagc gcgccagatt aaggatatat ggaaaaataa ccaacatcat
     2701 aaagatggcg gcgaacccta taaactcgca caacgccttg ccatgttagc ccatgaaatt
     2761 gacgcggtac ccgcctggaa ttgtaaaagc ggcaaagatc gtacagggat gatggattca
     2821 gaaatcaagc gagagatcat ttccttacat cagacccata tgttaagtgc gcctggtagt
     2881 cttccggata gcggtggaca gaaaattttc caaaaagtat tactgaatag cggtaacctg
     2941 gagattcaga aacaaaatac gggcggggcg ggaaacgcgg taatgaaaaa tttatcgcca
     3001 gaggtgctca atctttccta tcaaaaacga gttggggatg aaaatatttg gcagtcagta
     3061 aaaggcattt cttcattaat cacatcttaa ggatccaccg gatctagata actgatcata
     3121 atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc
     3181 ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat
     3241 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     3301 cattctagtt gtggtttgtc caaactcatc aatgtatctt aacgcgtaaa ttgtaagcgt
     3361 taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata
     3421 ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt
     3481 tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg
     3541 aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt
     3601 ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc
     3661 ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg
     3721 cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct
     3781 taatgcgccg ctacagggcg cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     3841 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     3901 ataaatgctt caataatatt gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg
     3961 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     4021 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     4081 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     4141 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     4201 ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga
     4261 ggaggctttt ttggaggcct aggcttttgc aaagatcgat caagagacag gatgaggatc
     4321 gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag
     4381 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg
     4441 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa
     4501 tgaactgcaa gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc
     4561 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc
     4621 ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga
     4681 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa
     4741 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct
     4801 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgagcat
     4861 gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt
     4921 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta
     4981 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga
     5041 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg
     5101 ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg
     5161 cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     5221 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     5281 ttcttcgccc accctagggg gaggctaact gaaacacgga aggagacaat accggaagga
     5341 acccgcgcta tgacggcaat aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt
     5401 cataaacgcg gggttcggtc ccagggctgg cactctgtcg ataccccacc gagaccccat
     5461 tggggccaat acgcccgcgt ttcttccttt tccccacccc accccccaag ttcgggtgaa
     5521 ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc aggttactca
     5581 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     5641 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     5701 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     5761 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     5821 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
     5881 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     5941 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     6001 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     6061 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     6121 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     6181 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     6241 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     6301 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     6361 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgt
//
