LOCUS       sequence-174779-        6070 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of KPTN.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6070)
  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 6070)
  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: pRK5-MYC-KPTN
FEATURES             Location/Qualifiers
     source          1..6070
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     581..605
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        815..833
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     815..832
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     846..865
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      950..959
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             959..988
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     regulatory      2357..2366
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2370..2504
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2407..2426)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2461..2480
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2573..2930
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2573..2764
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2596..2616)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2781..2916
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2950..2967)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2950..2966)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2959..2981)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3179..3634
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3266..3285)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3476..3497
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3651..3670)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3770..3792
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3830..3848)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3916..4020
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4021..4089,4090..4881)
                     /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     complement(4239..4258)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5052..5640
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5541..5560
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5794..5811
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5928..5949
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5964..5969,5970..5987,5988..5994)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6002..6018
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     6007..6029
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6026..6042
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6026..6042
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      601 agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagacac cgggaccgat
      661 ccagcctccg cggccgggaa cggtgcattg gaacgcggat tccccgtgcc aagagtgacg
      721 taagtaccgc ctatagagtc tataggccca cccccttggc ttcgttagaa cgcggctaca
      781 attaatacat aaccttatgt atcatacaca tacgatttag gtgacactat agaataacat
      841 ccactttgcc tttctctcca caggtgtcca ctcccaggtc caactgcacc tcggttctat
      901 cgattgaatt cccgaaccga cagtcggtct cttcaccaag gccattcgcg ccaccatgga
      961 gcaaaagctc atttctgagg aagatctcaa tggtggtggt ggtgggtcga cgatgatggg
     1021 ggaggcggcc gtggccgcgg ggccttgtcc gttgcgcgag gacagcttca cgcgcttctc
     1081 gtcgcagagc aatgtgtacg ggctggcagg cggcgccggc gggcgcgggg agctgctggc
     1141 cgccaccctt aaaggcaagg tgctcggctt ccgctaccaa gacctccgac agaaaatccg
     1201 gccagtggcc aaggagctgc agttcaacta cattcccgtg gatgcggaga ttgtctccat
     1261 cgacactttc aacaagtcac cccccaagcg gggtctggtt gtggggatca cgttcatcaa
     1321 ggattcaggg gacaagggca gccccttcct gaacatttac tgcgactacg agcccggctc
     1381 tgagtacaac cttgactcta ttgcccagag ctgcctgaac ctggagctcc agttcactcc
     1441 gttccagctg tgccatgcgg aggtccaggt cggggatcaa cttgagactg tgtttctctt
     1501 gagtgggaac gacccggcca ttcatctcta caaggagaac gaggggctgc atcagtttga
     1561 ggaacagccc gtggaaaacc tcttcccaga gctgacgaac ctgaccagta gcgtcctctg
     1621 gctggacgtc cacaacttcc ccggcacgtc ccggcgcctc tcagctctgg gctgtcagag
     1681 tggttatgtc cgtgtcgccc acgtggacca gcggagtcga gaggttctgc agatgtggtc
     1741 ggtcctgcag gacggtccca tctcccgagt gattgtgttc agcctctcgg ccgccaagga
     1801 gaccaaggac aggccactac aagatgagta cagcgtgctc gtggccagca tgttggagcc
     1861 agcagtggtg tatcgggacc tgctgaaccg gggtcttgaa gaccagcttc tcctgcccgg
     1921 cagtgaccag tttgacagcg tcctctgcag cctggtcacc gatgtggatt tggatgggcg
     1981 gccagaagtc ctggtggcca cctatggaca ggaactgctg tgttataagt accggggccc
     2041 agagtcgggg cttcctgagg cccagcacgg gttccatctg ctgtggcagc ggagcttctc
     2101 cagtcccctg ctggccatgg ctcacgtgga cctgaccggg gatgggctgc aggagcttgc
     2161 cgtggtctcc ctgaagggcg tgcacatcct gcagcacagc ctgattcagg cctcagagct
     2221 ggtcttgacc cggcttcgac atcaagtgga gcagaggaga cgtcggctac aggggttgga
     2281 ggacggggca ggtgcagggc ctgctgagaa tgcagcctct taagcggccg ctaagtaagt
     2341 aaggatcccc agcttggccg ccatggccca acttgtttat tgcagcttat aatggttaca
     2401 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     2461 gtggtttgtc caaactcatc aatgtatctt atcatgtctg gatcgggaat taattcggcg
     2521 cagcaccatg gcctgaaata acctctgaaa gaggaacttg gttaggtacc ttctgaggcg
     2581 gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag
     2641 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc
     2701 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag
     2761 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     2821 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc
     2881 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctgttaa
     2941 cagcttggca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca
     3001 acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg aagaggcccg
     3061 caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggcgcc tgatgcggta
     3121 ttttctcctt acgcatctgt gcggtatttc acaccgcata cgtcaaagca accatagtac
     3181 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
     3241 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     3301 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     3361 gctttacggc acctcgaccc caaaaaactt gatttgggtg atggttcacg tagtgggcca
     3421 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     3481 ctcttgttcc aaactggaac aacactcaac cctatctcgg gctattcttt tgatttataa
     3541 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     3601 gcgaatttta acaaaatatt aacgtttaca attttatggt gcactctcag tacaatctgc
     3661 tctgatgccg catagttaag ccagccccga cacccgccaa cacccgctga cgcgccctga
     3721 cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc
     3781 atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata
     3841 cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact
     3901 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     3961 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     4021 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     4081 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     4141 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     4201 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     4261 cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     4321 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     4381 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     4441 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     4501 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     4561 cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     4621 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     4681 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
     4741 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     4801 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     4861 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     4921 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     4981 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     5041 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     5101 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     5161 gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta
     5221 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     5281 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     5341 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     5401 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     5461 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     5521 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     5581 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     5641 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     5701 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
     5761 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
     5821 gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg
     5881 cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttag
     5941 ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga
     6001 attgtgagcg gataacaatt tcacacagga aacagctatg acatgattac gaattaattc
     6061 gagctcgccc
//
