LOCUS       sequence-174794-        8383 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of mios.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8383)
  AUTHORS   Wolfson RL, Chantranupong L, Wyant GA, Gu X, Orozco JM, Shen K, 
            Condon KJ, Petri S, Kedir J, Scaria SM, Abu-Remaileh M, Frankel WN, 
            Sabatini DM
  TITLE     KICSTOR recruits GATOR1 to the lysosome and is necessary for 
            nutrients to regulate mTORC1.
  JOURNAL   Nature. 2017 Feb 15. doi: 10.1038/nature21423.
  PUBMED    28199306
REFERENCE   2  (bases 1 to 8383)
  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: "Nature. 
            2017 Feb 15. doi: 10.1038/nature21423."
COMMENT     Sequence Label: KG371-GFP-mios
FEATURES             Location/Qualifiers
     source          1..8383
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..470
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    535..734
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             935..1351
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1281..1303
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1319..1335
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     CDS             4013..4030
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             4040..4063
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             4064..4087
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             join(4097..4099,4100..4102,4103..4816)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTATGITLGMDELYK"
     primer_bind     complement(4142..4163)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4403..4422)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4750..4771
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4961..4981)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4966..5295
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5146..5281
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5303..5701
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     LTR             5870..6339
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(6469..6486)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6640..7228)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6720..6739)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7388..8179,8180..8248))
                     /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     8011..8030
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8249..8353)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctgaatatgg gccaaacagg atatctgtgg taagcagttc ctgccccggc tcagggccaa
       61 gaacagatgg aacagctgaa tatgggccaa acaggatatc tgtggtaagc agttcctgcc
      121 ccggctcagg gccaagaaca gatggtcccc agatgcggtc cagccctcag cagtttctag
      181 agaaccatca gatgtttcca gggtgcccca aggacctgaa atgaccctgt gccttatttg
      241 aactaaccaa tcagttcgct tctcgcttct gttcgcgcgc ttctgctccc cgagctcaat
      301 aaaagagccc acaacccctc actcggggcg ccagtcctcc gattgactga gtcgcccggg
      361 tacccgtgta tccaataaac cctcttgcag ttgcatccga cttgtggtct cgctgttcct
      421 tgggagggtc tcctctgagt gattgactac ccgtcagcgg gggtctttca tttgggggct
      481 cgtccgggat cgggagaccc ctgcccaggg accaccgacc caccaccggg aggcaagctg
      541 gccagcaact tatctgtgtc tgtccgattg tctagtgtct atgactgatt ttatgcgcct
      601 gcgtcggtac tagttagcta actagctctg tatctggcgg acccgtggtg gaactgacga
      661 gttctgaaca cccggccgca accctgggag acgtcccagg gactttgggg gccgtttttg
      721 tggcccgacc tgaggaaggg agtcgatgtg gaatccgacc ccgtcaggat atgtggttct
      781 ggtaggagac gagaacctaa aacagttccc gcctccgtct gaatttttgc tttcggtttg
      841 gaaccgaagc cgcgcgtctt gtctgctgca gcgctgcagc atcgttctgt gttgtctctg
      901 tctgactgtg tttctgtatt tgtctgaaaa ttagggccag actgttacca ctcccttaag
      961 tttgacctta ggtcactgga aagatgtcga gcggatcgct cacaaccagt cggtagatgt
     1021 caagaagaga cgttgggtta ccttctgctc tgcagaatgg ccaaccttta acgtcggatg
     1081 gccgcgagac ggcaccttta accgagacct catcacccag gttaagatca aggtcttttc
     1141 acctggcccg catggacacc cagaccaggt cccctacatc gtgacctggg aagccttggc
     1201 ttttgacccc cctccctggg tcaagccctt tgtacaccct aagcctccgc ctcctcttcc
     1261 tccatccgcc ccgtctctcc cccttgaacc tcctcgttcg accccgcctc gatcctccct
     1321 ttatccagcc ctcactcctt ctctaggcgc cggccggatc ctcgaggcca ccatgtcggg
     1381 caccaaaccg gatattctgt gggcaccgca tcatgttgac cgcttcgttg tctgtgactc
     1441 ggaactgtcc ctgtaccatg tcgaaagtac cgtgaacagc gaactgaaag caggcagcct
     1501 gcgtctgtca gaagactcgg cggccaccct gctgagcatt aactctgata cgccgtatat
     1561 gaaatgtgtc gcgtggtatc tgaactacga tccggaatgc ctgctggccg ttggccaggc
     1621 aaatggtcgc gtggttctga ccagcctggg ccaagaccac aactcaaaat tcaaagatct
     1681 gatcggtaaa gaatttgtgc cgaaacatgc ccgtcagtgt aacaccctgg catggaatcc
     1741 gctggattct aactggctgg cagctggcct ggacaaacac cgcgcggatt tttctgtgct
     1801 gatttgggac atctgcagta aatatacccc ggatattgtt ccgatggaaa aagtcaaact
     1861 gtccgctggc gaaaccgaaa ccacgctgct ggttacgaaa ccgctgtacg aactgggtca
     1921 gaatgacgcc tgtctgtcac tgtgctggct gccgcgtgat caaaaactgc tgctggctgg
     1981 tatgcaccgt aatctggcga tttttgatct gcgcaacacc tcgcagaaaa tgttcgtgaa
     2041 taccaaagcg gttcaaggcg tgacggttga cccgtatttt catgatcgtg tcgctagctt
     2101 ctacgagggt caggtggcga tctgggatct gcgcaaattt gaaaaaccag tgctgaccct
     2161 gacggaacaa ccgaaaccgc tgaccaaagt cgcatggtgt ccgacccgta cgggcctgct
     2221 ggcaaccctg acgcgtgact ctaatattat ccgcctgtat gatatgcagc acaccccgac
     2281 gccgattggt gacgaaaccg aaccgacgat tatcgaacgc agtgtgcagc cgtgcgataa
     2341 ctacattgct tcgttcgcgt ggcatccgac cagccaaaac cgtatgatcg tcgtgacccc
     2401 gaatcgcacg atgagcgatt ttaccgtttt cgaacgtatt tcactggctt ggtcgccgat
     2461 cacgagcctg atgtgggcgt gtggtcgcca cctgtatgaa tgcaccgaag aagaaaacga
     2521 caattcgctg gaaaaagata ttgccacgaa aatgcgtctg cgcgcactga gccgttacgg
     2581 cctggacacc gaacaagtgt ggcgcaatca catcctggcc ggtaacgaag atccgcaact
     2641 gaaatccctg tggtataccc tgcattttat gaaacagtac acggaagata tggaccaaaa
     2701 atcgccgggc aataaaggta gcctggttta tgcgggcatt aaaagcatcg tgaaaagctc
     2761 tctgggtatg gttgaaagtt cccgtcataa ctggtctggc ctggataaac agagtgacat
     2821 tcaaaacctg aatgaagaac gcatcctggc cctgcagctg tgtggctgga ttaagaaagg
     2881 caccgatgtg gatgtgggcc cgtttctgaa cagcctggtg caagaaggtg aatgggaacg
     2941 cgcggcggca gttgcgctgt tcaacctgga tattcgtcgc gccattcaga tcctgaatga
     3001 aggcgcatca tcggaaaaag gtgatctgaa cctgaatgtt gtcgctatgg cgctgagtgg
     3061 ttataccgat gagaaaaaca gcctgtggcg tgaaatgtgt agtacgctgc gcctgcagct
     3121 gaacaatccg tacctgtgcg tgatgtttgc tttcctgacc tctgaaacgg gcagttatga
     3181 tggtgtcctg tacgaaaata aagttgcggt ccgtgaccgc gtggcctttg catgtaaatt
     3241 cctgtcagat acccagctga accgttatat cgaaaaactg acgaacgaaa tgaaagaagc
     3301 cggcaacctg gaaggtatcc tgctgaccgg cctgacgaaa gatggtgttg acctgatgga
     3361 aagctatgtc gatcgcaccg gcgacgtgca gacggcatcc tactgcatgc tgcaaggttc
     3421 accgctggat gttctgaaag acgaacgtgt ccagtattgg attgaaaact accgcaatct
     3481 gctggatgct tggcgttttt ggcacaaacg cgcggaattt gacatccatc gttctaaact
     3541 ggatccgagc tctaaaccgc tggcccaggt gtttgttagt tgtaacttct gcggcaaatc
     3601 aatttcgtat agctgctctg cagtgccgca tcagggtcgc ggttttagcc aatacggcgt
     3661 tagtggttcc ccgaccaaat ccaaagtcac gtcatgtccg ggttgccgta aaccgctgcc
     3721 gcgttgtgca ctgtgcctga tcaatatggg taccccggtg tcttcctgtc cgggcggtac
     3781 gaaatcggat gaaaaagttg acctgagcaa agataaaaaa ctggcccagt ttaacaattg
     3841 gttcacctgg tgtcacaact gccgtcatgg cggtcacgca ggtcatatgc tgtcctggtt
     3901 tcgcgatcat gctgaatgcc cggtgtcagc gtgcacctgt aaatgcatgc aactggatac
     3961 gacgggtaat ctggtcccgg cggaaaccgt tcaaccgggc cgcggcagca gccatcatca
     4021 tcatcatcac agcagcggcc tggaagttct gttccagggg cccctggaag ttctgttcca
     4081 ggggccctcc ggagggatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat
     4141 cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga
     4201 gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc
     4261 cgtgccctgg cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta
     4321 ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca
     4381 ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt
     4441 cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg
     4501 caacatcctg gggcacaagc tggagtacaa ctacaacagc cacaacgtct atatcatggc
     4561 cgacaagcag aagaacggca tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg
     4621 cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct
     4681 gctgcccgac aaccactacc tgagcaccca gtccgccctg agcaaagacc ccaacgagaa
     4741 gcgcgatcac atggtcctgc tggagttcgt gaccgccacc gggatcactc ttggcatgga
     4801 cgagctgtac aagtaaagat ccaccggatc tagataactg atcataatca gccataccac
     4861 atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca
     4921 taaaatgaat gcaattcgcc agcacagtgg tcgaccctgt ggaatgtgtg tcagttaggg
     4981 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     5041 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     5101 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     5161 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     5221 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     5281 ctaggctttt gcaaaaagct tcatggccaa gcctttgtct caagaagaat ccaccctcat
     5341 tgaaagagca acggctacaa tcaacagcat ccccatctct gaagactaca gcgtcgccag
     5401 cgcagctctc tctagcgacg gccgcatctt cactggtgtc aatgtatatc attttactgg
     5461 gggaccttgt gcagaactcg tggtgctggg cactgctgct gctgcggcag ctggcaacct
     5521 gacttgtatc gtcgcgatcg gaaatgagaa caggggcatc ttgagcccct gcggacggtg
     5581 ccgacaggtg cttctcgatc tgcatcctgg gatcaaagcc atagtgaagg acagtgatgg
     5641 acagccgacg gcagttggga ttcgtgaatt gctgccctct ggttatgtgt gggagggcta
     5701 agatcgataa aataaaagat tttatttagt ctccagaaaa aggggggaat gaaagacccc
     5761 acctgtaggt ttggcaagct agcttaagta acgccatttt gcaaggcatg gaaaaataca
     5821 taactgagaa tagagaagtt cagatcaagg tcaggaacag atggaacagc tgaatatggg
     5881 ccaaacagga tatctgtggt aagcagttcc tgccccggct cagggccaag aacagatgga
     5941 acagctgaat atgggccaaa caggatatct gtggtaagca gttcctgccc cggctcaggg
     6001 ccaagaacag atggtcccca gatgcggtcc agccctcagc agtttctaga gaaccatcag
     6061 atgtttccag ggtgccccaa ggacctgaaa tgaccctgtg ccttatttga actaaccaat
     6121 cagttcgctt ctcgcttctg ttcgcgcgct tctgctcccc gagctcaata aaagagccca
     6181 caacccctca ctcggggcgc cagtcctccg attgactgag tcgcccgggt acccgtgtat
     6241 ccaataaacc ctcttgcagt tgcatccgac ttgtggtctc gctgttcctt gggagggtct
     6301 cctctgagtg attgactacc cgtcagcggg ggtctttcac atgcagcatg tatcaaaatt
     6361 aatttggttt tttttcttaa gtatttacat taaatggcca tagttgcatt aatgaatcgg
     6421 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     6481 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     6541 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     6601 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     6661 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     6721 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     6781 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     6841 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     6901 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     6961 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     7021 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     7081 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     7141 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     7201 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     7261 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     7321 cttcacctag atccttttgc ggccgcaaat caatctaaag tatatatgag taaacttggt
     7381 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     7441 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     7501 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     7561 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     7621 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     7681 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     7741 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     7801 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     7861 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     7921 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     7981 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     8041 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     8101 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     8161 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     8221 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     8281 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     8341 taggggttcc gcgcacattt ccccgaaaag tgccacctgc agc
//
