LOCUS       sequence-174784-        6109 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 6109)
  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 6109)
  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-HA-ITFG2
FEATURES             Location/Qualifiers
     source          1..6109
                     /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      2396..2405
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2409..2543
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2446..2465)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2500..2519
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2612..2969
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2612..2803
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2635..2655)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2820..2955
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2989..3006)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2989..3005)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2998..3020)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3218..3673
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3305..3324)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3515..3536
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3690..3709)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3809..3831
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3869..3887)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3955..4059
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4060..4128,4129..4920)
                     /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(4278..4297)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5091..5679
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5580..5599
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5833..5850
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5967..5988
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6003..6008,6009..6026,6027..6033)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6041..6057
                     /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     6046..6068
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6065..6081
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6065..6081
                     /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 gaggtcagtt agctacgtgc agcgcgtggc gctggagttc agcgggagcc tcttcccgca
     1081 cgcaatctgc ctcggagacg ttgataacga tacgttaaat gaactggtgg tgggagacac
     1141 cagcgggaag gtgtctgtgt ataaaaatga tgacagtcgg ccatggctca cctgttcctg
     1201 ccagggaatg ctgacttgcg ttggggttgg agacgtgtgt aataaaggaa agaacctgtt
     1261 ggtggcagtg agtgctgaag gctggtttca tttgtttgac ctgacacctg ccaaggtgtt
     1321 ggatgcttct gggcaccacg agacactaat cggagaggag cagcgtccag tcttcaagca
     1381 gcacatccct gccaacacca aggtcatgct gatcagcgac atcgatggag atgggtgtcg
     1441 tgagctggtg gtgggctaca cagaccgtgt ggtgcgagct ttccgctggg aggagctagg
     1501 tgagggtcct gaacatctga cagggcagct ggtgtccctc aagaaatgga tgctggaggg
     1561 tcaggtggac agcctctcag tgactctggg gccactgggt cttcctgaac tgatggtgtc
     1621 tcagccaggt tgtgcgtatg caattctact gtgtacctgg aaaaaggaca ctgggtcccc
     1681 tcctgcctct gaagggccca cggatggtag tagggagacc ccagctgccc gagacgtggt
     1741 gctgcaccag acatctggcc gtatccacaa caagaatgtc tccactcacc taattggcaa
     1801 catcaaacaa ggccacggca ctgagagtag tggctctggc ctctttgccc tgtgcaccct
     1861 ggatgggaca ctgaagctca tggaagaaat ggaagaagca gacaagctgc tgtggtcagt
     1921 gcaggtggat caccagctct ttgccctgga gaaactggat gtcaccggca acgggcatga
     1981 ggaggtagtt gcatgcgcct gggatggaca gacatatatc attgatcaca accgcaccgt
     2041 cgtccgcttc caagtggatg aaaatatccg tgccttctgt gcaggcctgt acgcctgcaa
     2101 agagggccgc aacagcccct gcctcgtata tgtcactttc aaccagaaga tctatgtgta
     2161 ctgggaggtg cagctggagc ggatggagtc taccaatctg gtgaaactgc tggagaccaa
     2221 gccggagtac cacagcctgc tgcaggagct gggcgtggat cctgacgacc tccctgtgac
     2281 tcgtgccctg cttcaccaaa cgctctacca tccagaccag ccaccacagt gtgctccctc
     2341 aagcctccag gatcccacct aggcggccgc taagtaagta aggatcccca gcttggccgc
     2401 catggcccaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     2461 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     2521 atgtatctta tcatgtctgg atcgggaatt aattcggcgc agcaccatgg cctgaaataa
     2581 cctctgaaag aggaacttgg ttaggtacct tctgaggcgg aaagaaccag ctgtggaatg
     2641 tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     2701 tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa
     2761 gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta actccgccca
     2821 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
     2881 ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag
     2941 gcttttttgg aggcctaggc ttttgcaaaa agctgttaac agcttggcac tggccgtcgt
     3001 tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca
     3061 tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca
     3121 gttgcgcagc ctgaatggcg aatggcgcct gatgcggtat tttctcctta cgcatctgtg
     3181 cggtatttca caccgcatac gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta
     3241 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg
     3301 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     3361 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     3421 aaaaaacttg atttgggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     3481 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
     3541 acactcaacc ctatctcggg ctattctttt gatttataag ggattttgcc gatttcggcc
     3601 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta
     3661 acgtttacaa ttttatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
     3721 cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca
     3781 tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg
     3841 tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt ataggttaat
     3901 gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga
     3961 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     4021 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     4081 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     4141 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     4201 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     4261 agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     4321 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     4381 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     4441 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     4501 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     4561 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     4621 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     4681 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     4741 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     4801 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     4861 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     4921 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     4981 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     5041 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     5101 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     5161 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     5221 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     5281 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     5341 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     5401 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     5461 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     5521 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     5581 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     5641 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     5701 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     5761 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     5821 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg
     5881 cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg
     5941 aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta ggcaccccag
     6001 gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt
     6061 cacacaggaa acagctatga catgattacg aattaattcg agctcgccc
//
