LOCUS       sequence-174181-        5693 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  GFP-tagged mSin1 (isoform 5).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5693)
  AUTHORS   Ebner M, Sinkovics B, Szczygiel M, Ribeiro DW, Yudushkin I
  TITLE     Localization of mTORC2 activity inside cells.
  JOURNAL   J Cell Biol. 2017 Jan 31. pii: jcb.201610060. doi: 
            10.1083/jcb.201610060.
  PUBMED    28143890
REFERENCE   2  (bases 1 to 5693)
  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: "J Cell 
            Biol. 2017 Jan 31. pii: jcb.201610060. doi: 10.1083/jcb.201610060."
COMMENT     Sequence Label: N1-mSin1.5-GFP
FEATURES             Location/Qualifiers
     source          1..5693
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      136..271
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             320..1114
                     /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             320..1114
                     /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(374..393)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     984..1003
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1302..1321)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1346..1393
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      1722..2310
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2211..2230
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        2485..2788
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2789..2992
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2943..2963
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             join(4066..4068,4069..4782)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(4066..4068,4069..4779)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(4108..4129)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4369..4388)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4716..4737
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    4905..5026
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4942..4961)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4996..5015
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5033..5488)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5170..5191)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5382..5401
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5515..5619
                     /gene="bla"
                     /label=AmpR promoter
     promoter        join(5621..5693,1..285)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        join(5621..5693,1..119)
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5644..5664)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
ORIGIN
        1 agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc
       61 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg
      121 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
      181 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc
      241 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaagatcg atcaagagac
      301 aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc
      361 ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc
      421 cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc
      481 cggtgccctg aatgaactgc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg
      541 cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt
      601 gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc
      661 catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga
      721 ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga
      781 tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct
      841 caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc
      901 gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt
      961 ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg
     1021 cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat
     1081 cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc
     1141 gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa
     1201 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat
     1261 ctcatgctgg agttcttcgc ccaccctagg gggaggctaa ctgaaacacg gaaggagaca
     1321 ataccggaag gaacccgcgc tatgacggca ataaaaagac agaataaaac gcacggtgtt
     1381 gggtcgtttg ttcataaacg cggggttcgg tcccagggct ggcactctgt cgatacccca
     1441 ccgagacccc attggggcca atacgcccgc gtttcttcct tttccccacc ccacccccca
     1501 agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc tgccatagcc
     1561 tcaggttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
     1621 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     1681 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     1741 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     1801 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     1861 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     1921 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     1981 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     2041 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     2101 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     2161 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     2221 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     2281 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     2341 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     2401 gataaccgta ttaccgccat gcattagtta ttaatagtaa tcaattacgg ggtcattagt
     2461 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
     2521 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
     2581 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
     2641 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
     2701 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
     2761 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg
     2821 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
     2881 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt
     2941 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctggtttagt
     3001 gaaccgtcag atccgctagc gctaccggac tcagatctcg agctcaagct tcgaattccc
     3061 accatggcct tcttggacaa tccaactatc attctagctc atattcgaca gtcacatgtg
     3121 accagtgatg acacgggaat gtgtgagatg gttctcattg atcatgatgt tgacctagag
     3181 aagattcatc ctccttcaat gcctggagac agtgggtcag aaattcaggg aagcaatggt
     3241 gagactcagg gctatgtata tgcccagtca gtcgatatta cctcaagttg ggactttggt
     3301 attagaagac gctcaaacac agctcaaaga ttagaacgac tccgaaaaga gagacaaaac
     3361 cagatcaaat gcaaaaatat tcagtggaaa gaaagaaatt ctaagcaatc agcccaggag
     3421 ttaaagtcac tgtttgaaaa aaaatctctc aaagagaagc ctccaatttc tgggaagcag
     3481 tcgatattat ctgtacgcct agaacagtgc cctctgcagc tgaataaccc ttttaacgag
     3541 tattccaaat ttgatggcaa gggtcatgta ggtacaacag caaccaagaa gatcgatgtc
     3601 tacctccctc tgcactcgag ccaggacaga ctgctgccaa tgaccgtggt gacaatggcc
     3661 agcgccaggg tgcaggacct gatcgggctc atctgctggc agtatacaag cgaaggacgg
     3721 gagccgaagc tcaatgacaa tgtcagtgcc tactgcctgc atattgctga ggatgatggg
     3781 gaggtggaca ccgatttccc cccgctggat tccaatgagc ccattcataa gtttggcttc
     3841 agtactttgg ccctggttga aaagtactca tctcctggtc tgacatccaa agagtcactc
     3901 tttgttcgaa taaatgctgc tcatggattc tcccttattc aggtggacaa cacaaaggtt
     3961 accatgaagg aaatcttact gaaggcagtg aagcgaagaa aaggatccca gaaagtttca
     4021 ggtgcttgtg acccgcgggc ccgggatcca ccggtcgcaa ccttggtgag caagggcgag
     4081 gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac
     4141 aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag
     4201 ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc
     4261 tacggcgtgc agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag
     4321 tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac
     4381 tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg
     4441 aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac
     4501 aacagccaca acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc
     4561 aagatccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac
     4621 acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc
     4681 gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc
     4741 gccgccggga tcactctcgg catggacgag ctgtacaagt aaagcggccg cgactctaga
     4801 tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc
     4861 tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag
     4921 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     4981 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttaaggc gtaaattgta
     5041 agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac
     5101 caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg
     5161 agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa
     5221 gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt
     5281 tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt
     5341 agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga
     5401 gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc
     5461 gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga
     5521 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     5581 ccctgataaa tgcttcaata atattgaaaa aggaagagtc ctgaggcgga aagaaccagc
     5641 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggc
//
