LOCUS       sequence-173584-        4777 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Akt pseudosubstrate tagged with mCherry.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4777)
  AUTHORS   Ebner M, Lucic I, Leonard TA, Yudushkin I
  TITLE     PI(3,4,5)P3 Engagement Restricts Akt Activity to Cellular Membranes.
  JOURNAL   Mol Cell. 2017 Feb 2;65(3):416-431.e6. doi: 
            10.1016/j.molcel.2016.12.028.
  PUBMED    28157504
REFERENCE   2  (bases 1 to 4777)
  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.molcel.2016.12"; journalName: "Mol Cell"; date: 
            "2017-02-2- 2"; volume: "65"; issue: "3"; pages: "416-431.e6"
COMMENT     Sequence Label: pmCherry-PS
FEATURES             Location/Qualifiers
     source          1..4777
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      11..599
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     500..519
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        774..1077
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1078..1281
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1232..1252
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1320..1329
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1326..2033
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1471..1489)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1739..1758
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    2278..2399
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2315..2334)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2369..2388
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2406..2861)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2543..2564)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2755..2774
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2888..2992
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2994..3351
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2994..3185
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3017..3037)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3202..3337
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3386..4180
                     /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             3386..4180
                     /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(3440..3459)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4050..4069
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4368..4387)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4412..4459
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
ORIGIN
        1 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
       61 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
      121 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
      181 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
      241 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
      301 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
      361 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
      421 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
      481 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
      541 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
      601 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
      661 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccatg cattagttat
      721 taatagtaat caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca
      781 taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca
      841 ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg
      901 gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg
      961 ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc
     1021 ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg
     1081 atgcggtttt ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca
     1141 agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt
     1201 ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg
     1261 gaggtctata taagcagagc tggtttagtg aaccgtcaga tccgctagcg ctaccggtcg
     1321 ccaccatggt gagcaagggc gaggaggata acatggccat catcaaggag ttcatgcgct
     1381 tcaaggtgca catggagggc tccgtgaacg gccacgagtt cgagatcgag ggcgagggcg
     1441 agggccgccc ctacgagggc acccagaccg ccaagctgaa ggtgaccaag ggtggccccc
     1501 tgcccttcgc ctgggacatc ctgtcccctc agttcatgta cggctccaag gcctacgtga
     1561 agcaccccgc cgacatcccc gactacttga agctgtcctt ccccgagggc ttcaagtggg
     1621 agcgcgtgat gaacttcgag gacggcggcg tggtgaccgt gacccaggac tcctccctgc
     1681 aggacggcga gttcatctac aaggtgaagc tgcgcggcac caacttcccc tccgacggcc
     1741 ccgtaatgca gaagaagacc atgggctggg aggcctcctc cgagcggatg taccccgagg
     1801 acggcgccct gaagggcgag atcaagcaga ggctgaagct gaaggacggc ggccactacg
     1861 acgctgaggt caagaccacc tacaaggcca agaagcccgt gcagctgccc ggcgcctaca
     1921 acgtcaacat caagttggac atcacctccc acaacgagga ctacaccatc gtggaacagt
     1981 acgaacgcgc cgagggccgc cactccaccg gcggcatgga cgagctgtac aagtccggac
     2041 tcagatctcg agctgttgaa ctagatcctg agtttgaacc tcgtgccaga gagcgtgctt
     2101 acgccttcgg acactaagaa ttctgcagtc gacggtaccg cgggcccggg atccaccgga
     2161 tctagataac tgatcataat cagccatacc acatttgtag aggttttact tgctttaaaa
     2221 aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt tgttgttaac
     2281 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     2341 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttaa
     2401 cgcgtaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
     2461 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
     2521 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
     2581 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
     2641 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
     2701 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
     2761 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
     2821 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcaggtggca cttttcgggg
     2881 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     2941 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtcctgaggc
     3001 ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     3061 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     3121 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     3181 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     3241 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag
     3301 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa agatcgatca
     3361 agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc
     3421 ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc
     3481 tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga
     3541 cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt ggctggccac
     3601 gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct
     3661 gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa
     3721 agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc
     3781 attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct
     3841 tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc
     3901 caggctcaag gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg
     3961 cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct
     4021 gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct
     4081 tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca
     4141 gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa
     4201 atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc
     4261 tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc
     4321 ggggatctca tgctggagtt cttcgcccac cctaggggga ggctaactga aacacggaag
     4381 gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa taaaacgcac
     4441 ggtgttgggt cgtttgttca taaacgcggg gttcggtccc agggctggca ctctgtcgat
     4501 accccaccga gaccccattg gggccaatac gcccgcgttt cttccttttc cccaccccac
     4561 cccccaagtt cgggtgaagg cccagggctc gcagccaacg tcggggcggc aggccctgcc
     4621 atagcctcag gttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     4681 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     4741 tcgttccact gagcgtcaga ccccgtagaa aagatca
//
