LOCUS       sequence-174782-        6056 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of ITFG2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6056)
  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 6056)
  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-FLAG-ITFG2
FEATURES             Location/Qualifiers
     source          1..6056
                     /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"
     CDS             924..947
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      2343..2352
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2356..2490
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2393..2412)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2447..2466
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2559..2916
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2559..2750
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2582..2602)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2767..2902
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2936..2953)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2936..2952)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2945..2967)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3165..3620
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3252..3271)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3462..3483
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3637..3656)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3756..3778
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3816..3834)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3902..4006
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4007..4075,4076..4867)
                     /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(4225..4244)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5038..5626
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5527..5546
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5780..5797
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5914..5935
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5950..5955,5956..5973,5974..5980)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5988..6004
                     /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     5993..6015
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6012..6028
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6012..6028
                     /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 caccatggaa atggactaca aggatgacga tgacaagggt ggtggtggtg
      961 gttcgacgat gaggtcagtt agctacgtgc agcgcgtggc gctggagttc agcgggagcc
     1021 tcttcccgca cgcaatctgc ctcggagacg ttgataacga tacgttaaat gaactggtgg
     1081 tgggagacac cagcgggaag gtgtctgtgt ataaaaatga tgacagtcgg ccatggctca
     1141 cctgttcctg ccagggaatg ctgacttgcg ttggggttgg agacgtgtgt aataaaggaa
     1201 agaacctgtt ggtggcagtg agtgctgaag gctggtttca tttgtttgac ctgacacctg
     1261 ccaaggtgtt ggatgcttct gggcaccacg agacactaat cggagaggag cagcgtccag
     1321 tcttcaagca gcacatccct gccaacacca aggtcatgct gatcagcgac atcgatggag
     1381 atgggtgtcg tgagctggtg gtgggctaca cagaccgtgt ggtgcgagct ttccgctggg
     1441 aggagctagg tgagggtcct gaacatctga cagggcagct ggtgtccctc aagaaatgga
     1501 tgctggaggg tcaggtggac agcctctcag tgactctggg gccactgggt cttcctgaac
     1561 tgatggtgtc tcagccaggt tgtgcgtatg caattctact gtgtacctgg aaaaaggaca
     1621 ctgggtcccc tcctgcctct gaagggccca cggatggtag tagggagacc ccagctgccc
     1681 gagacgtggt gctgcaccag acatctggcc gtatccacaa caagaatgtc tccactcacc
     1741 taattggcaa catcaaacaa ggccacggca ctgagagtag tggctctggc ctctttgccc
     1801 tgtgcaccct ggatgggaca ctgaagctca tggaagaaat ggaagaagca gacaagctgc
     1861 tgtggtcagt gcaggtggat caccagctct ttgccctgga gaaactggat gtcaccggca
     1921 acgggcatga ggaggtagtt gcatgcgcct gggatggaca gacatatatc attgatcaca
     1981 accgcaccgt cgtccgcttc caagtggatg aaaatatccg tgccttctgt gcaggcctgt
     2041 acgcctgcaa agagggccgc aacagcccct gcctcgtata tgtcactttc aaccagaaga
     2101 tctatgtgta ctgggaggtg cagctggagc ggatggagtc taccaatctg gtgaaactgc
     2161 tggagaccaa gccggagtac cacagcctgc tgcaggagct gggcgtggat cctgacgacc
     2221 tccctgtgac tcgtgccctg cttcaccaaa cgctctacca tccagaccag ccaccacagt
     2281 gtgctccctc aagcctccag gatcccacct aggcggccaa gtaagtaagg atccccagct
     2341 tggccgccat ggcccaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     2401 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     2461 ctcatcaatg tatcttatca tgtctggatc gggaattaat tcggcgcagc accatggcct
     2521 gaaataacct ctgaaagagg aacttggtta ggtaccttct gaggcggaaa gaaccagctg
     2581 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     2641 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     2701 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     2761 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     2821 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag
     2881 tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tgttaacagc ttggcactgg
     2941 ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg
     3001 cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt
     3061 cccaacagtt gcgcagcctg aatggcgaat ggcgcctgat gcggtatttt ctccttacgc
     3121 atctgtgcgg tatttcacac cgcatacgtc aaagcaacca tagtacgcgc cctgtagcgg
     3181 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     3241 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     3301 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     3361 cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt gggccatcgc cctgatagac
     3421 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     3481 tggaacaaca ctcaacccta tctcgggcta ttcttttgat ttataaggga ttttgccgat
     3541 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa
     3601 aatattaacg tttacaattt tatggtgcac tctcagtaca atctgctctg atgccgcata
     3661 gttaagccag ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct
     3721 cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt
     3781 ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc gtgatacgcc tatttttata
     3841 ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc ggggaaatgt
     3901 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
     3961 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
     4021 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
     4081 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
     4141 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
     4201 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
     4261 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
     4321 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
     4381 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     4441 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     4501 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     4561 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     4621 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     4681 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     4741 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     4801 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     4861 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     4921 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     4981 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     5041 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     5101 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     5161 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     5221 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     5281 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     5341 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     5401 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     5461 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     5521 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     5581 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     5641 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     5701 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     5761 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg
     5821 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
     5881 cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca ctcattaggc
     5941 accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg tgagcggata
     6001 acaatttcac acaggaaaca gctatgacat gattacgaat taattcgagc tcgccc
//
