LOCUS       sequence-194373-        5965 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of HA-RagC.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5965)
  AUTHORS   Shen K, Choe A, Sabatini DM
  TITLE     Intersubunit Crosstalk in the Rag GTPase Heterodimer Enables mTORC1 
            to Respond Rapidly to Amino Acid Availability.
  JOURNAL   Mol Cell. 2017 Oct 17. pii: S1097-2765(17)30704-9. doi: 
            10.1016/j.molcel.2017.09.026.
  PUBMED    29056322
REFERENCE   2  (bases 1 to 5965)
  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: "Mol Cell. 
            2017 Oct 17. pii: S1097-2765(17)30704-9. doi: 
            10.1016/j.molcel.2017.09.026."
COMMENT     Sequence Label: pRK5 HA-RagC
FEATURES             Location/Qualifiers
     source          1..5965
                     /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             962..988
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     regulatory      2252..2261
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2265..2399
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2302..2321)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2356..2375
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2468..2825
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2468..2659
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2491..2511)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2676..2811
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2845..2862)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2845..2861)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2854..2876)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3074..3529
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3161..3180)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3371..3392
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3546..3565)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3665..3687
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3725..3743)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3811..3915
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3916..3984,3985..4776)
                     /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(4134..4153)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4947..5535
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5436..5455
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5689..5706
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5823..5844
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5859..5864,5865..5882,5883..5889)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5897..5913
                     /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     5902..5924
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5921..5937
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5921..5937
                     /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 ccaccatggg
      961 ctacccctat gatgtgcctg actacgcaga tctcaatggt ggtggtggtg ggtcgacgat
     1021 gtccctgcag tacggggcgg aggagacgcc cctcgccggc agttacggcg cggccgattc
     1081 gtttccaaag gacttcggct acggcgtgga ggaggaggaa gaggaggcgg cggcggcggg
     1141 cggaggggtt ggggcagggg caggcggtgg ctgtggtccg gggggcgctg acagctccaa
     1201 gccgaggatt ctgctcatgg gactccggcg cagcggcaag tcctccatcc agaaggtggt
     1261 gtttcataag atgtcaccca acgagaccct ctttttggaa agtaccaaca agatttataa
     1321 ggatgacatt tccaatagct cctttgtgaa tttccagata tgggattttc ctgggcaaat
     1381 ggactttttt gacccaacct ttgactatga gatgatcttc aggggaacag gagcattgat
     1441 atacgtcatt gacgcacagg atgactacat ggaggcttta acaagacttc acattactgt
     1501 ttctaaagcc tacaaagtta acccagacat gaattttgag gtttttattc acaaagttga
     1561 tggtctgtct gatgatcaca aaatagaaac acagagggac attcatcaaa gggccaatga
     1621 tgaccttgca gatgctgggc tagaaaaact ccatcttagc ttttatctga ctagtatcta
     1681 tgaccattca atatttgaag cctttagtaa ggtggtgcag aaactcattc cacaactgcc
     1741 gaccttggaa aacctattaa atatctttat atcaaattca ggtattgaaa aagcttttct
     1801 ctttgatgtt gtcagcaaaa tctacattgc aacagacagt tcccctgtgg atatgcaatc
     1861 ttatgaactt tgctgtgaca tgatcgatgt tgtaattgat gtgtcttgta tatatgggtt
     1921 aaaggaagat ggaagtggaa gtgcttatga caaagaatct atggcaatta tcaagctgaa
     1981 taatacaact gtcctttatt taaaggaggt gactaaattt ttggcactgg tctgcattct
     2041 aagggaagaa agctttgaaa gaaaaggttt aatagactac aacttccact gtttccgaaa
     2101 agctattcat gaggtttttg aggtgggtgt gacttctcac aggagctgtg gtcaccagac
     2161 tagtgcctcc agtctgaaag cgctgacaca caatggcacg ccacgaaacg ccatctaggc
     2221 ggccgctaag taagtaagga tccccagctt ggccgccatg gcccaacttg tttattgcag
     2281 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     2341 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctggatcg
     2401 ggaattaatt cggcgcagca ccatggcctg aaataacctc tgaaagagga acttggttag
     2461 gtaccttctg aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt
     2521 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     2581 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     2641 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     2701 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     2761 cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     2821 gcaaaaagct gttaacagct tggcactggc cgtcgtttta caacgtcgtg actgggaaaa
     2881 ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa
     2941 tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg
     3001 gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtca
     3061 aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     3121 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     3181 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     3241 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgattt gggtgatggt
     3301 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     3361 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcgggctat
     3421 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     3481 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt atggtgcact
     3541 ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc
     3601 gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc
     3661 gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga
     3721 aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag
     3781 acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     3841 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     3901 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     3961 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     4021 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     4081 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     4141 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat
     4201 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     4261 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     4321 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     4381 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     4441 cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa
     4501 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     4561 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     4621 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     4681 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     4741 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     4801 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     4861 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     4921 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     4981 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     5041 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     5101 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     5161 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     5221 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     5281 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     5341 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     5401 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     5461 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     5521 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     5581 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     5641 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
     5701 agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt
     5761 cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca
     5821 attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct
     5881 cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag ctatgacatg
     5941 attacgaatt aattcgagct cgccc
//
