LOCUS       sequence-472456-        8275 bp    DNA     circular SYN 13-MAY-2025
DEFINITION  Expression of FL human Beclin 1 and FL human ATG14L in mammalian 
            cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8275)
  AUTHORS   Ohashi Y, Tremel S, Masson GR, McGinney L, Boulanger J, 
            Rostislavleva K, Johnson CM, Niewczas I, Clark J, Williams RL
  TITLE     Membrane characteristics tune activities of endosomal and autophagic
            human VPS34 complexes.
  JOURNAL   Elife. 2020 Jun 30;9:e58281. doi: 10.7554/eLife.58281.
  PUBMED    32602837
REFERENCE   2  (bases 1 to 8275)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Elife."; 
            date: "2020-06-30"; volume: "9:e58281. doi"; pages: " 
            10.7554/eLife.58281"
COMMENT     Sequence Label: pYO1101
FEATURES             Location/Qualifiers
     source          1..8275
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        2..381
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        382..585
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     536..556
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    587..605
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    608..626
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     636..660
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     primer_bind     complement(2147..2164)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2153..2377
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     intron          2630..3646
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     3570..3592
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     3654..3673
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     complement(5297..5316)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    5362..5417
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(5416..5435)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(5778..5794)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5778..5794)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5791..5813)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5802..5818
                     /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)."
     promoter        complement(join(5826..5832,5833..5850,5851..5856))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5871..5892
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5951..6146
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5997..6132
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    6152..6286
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6189..6208)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6243..6262
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6354..6371)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6525..7113)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6605..6624)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7284..8075,8076..8144))
                     /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     7907..7926
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8145..8249)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt agttcatagc
       61 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc
      121 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg
      181 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
      241 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc
      301 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta
      361 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag
      421 cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt
      481 tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa
      541 atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctctccc tatcagtgat
      601 agagatctcc ctatcagtga tagagatcgt cgacgagctc gtttagtgaa ccgtcagatc
      661 gcctggagac gccatccacg ctgttttgac ctccatagaa gacaccggga ccgatccagc
      721 ctccggactc tagcgtttaa acttaagctt ggtaccatgg aagggtctaa gacgtccaac
      781 aacagcacca tgcaggtgag cttcgtgtgc cagcgctgca gccagcccct gaaactggac
      841 acgagtttca agatcctgga ccgtgtcacc atccaggaac tcacagctcc attacttacc
      901 acagcccagg cgaaaccagg agagacccag gaggaagaga ctaactcagg agaggagcca
      961 tttattgaaa ctcctcgcca ggatggtgtc tctcgcagat tcatcccccc agccaggatg
     1021 atgtccacag aaagtgccaa cagcttcact ctgattgggg aggcatctga tggcggcacc
     1081 atggagaacc tcagccgaag actgaaggtc actggggacc tttttgacat catgtcgggc
     1141 cagacagatg tggatcaccc actctgtgag gaatgcacag atactctttt agaccagctg
     1201 gacactcagc tcaacgtcac tgaaaatgag tgtcagaact acaaacgctg tttggagatc
     1261 ttagagcaaa tgaatgagga tgacagtgaa cagttacaga tggagctaaa ggagctggca
     1321 ctagaggagg agaggctgat ccaggagctg gaagacgtgg aaaagaaccg caagatagtg
     1381 gcagaaaatc tcgagaaggt ccaggctgag gctgagagac tggatcagga ggaagctcag
     1441 tatcagagag aatacagtga atttaaacga cagcagctgg agctggatga tgagctgaag
     1501 agtgttgaaa accagatgcg ttatgcccag acgcagctgg ataagctgaa gaaaaccaac
     1561 gtctttaatg caaccttcca catctggcac agtggacagt ttggcacaat caataacttc
     1621 aggctgggtc gcctgcccag tgttcccgtg gaatggaatg agattaatgc tgcttggggc
     1681 cagactgtgt tgctgctcca tgctctggcc aataagatgg gtctgaaatt tcagagatac
     1741 cgacttgttc cttacggaaa ccattcatat ctggagtctc tgacagacaa atctaaggag
     1801 ctgccgttat actgttctgg ggggttgcgg tttttctggg acaacaagtt tgaccatgca
     1861 atggtggctt tcctggactg tgtgcagcag ttcaaagaag aggttgagaa aggcgagaca
     1921 cgtttttgtc ttccctacag gatggatgtg gagaaaggca agattgaaga cacaggaggc
     1981 agtggcggct cctattccat caaaacccag tttaactctg aggagcagtg gacaaaagct
     2041 ctcaagttca tgctgacgaa tctgaagtgg ggtctggctt gggtgtcctc tcaattttat
     2101 aacaaatgag tcgagtctag agggcccgtt taaacccgct gatcagcctc gactgtgcct
     2161 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt
     2221 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg
     2281 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac
     2341 aatagcaggc atgctgggga tgcggtgggc tctatggtat taatagtaat caattacggg
     2401 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
     2461 gcctggctga ccgcccaacg acccccgccc attgacgtca ataaggcggg gcggggcgag
     2521 gcggagaggt gcggcggcag ccaatcagag cggcgcgctc cgaaagtttc cttttatggc
     2581 gaggcggcgg cggcggcggc cctataaaaa gcgaagcgcg cggcgggcgg gagtcgctgc
     2641 gcgctgcctt cgccccgtgc cccgctccgc cgccgcctcg cgccgcccgc cccggctctg
     2701 actgaccgcg ttactcccac aggtgagcgg gcgggacggc ccttctcctc cgggctgtaa
     2761 ttagcgcttg gtttaatgac ggcttgtttc ttttctgtgg ctgcgtgaaa gccttgaggg
     2821 gctccgggag ggccctttgt gcggggggag cggctcgggg ggtgcgtgcg tgtgtgtgtg
     2881 cgtggggagc gccgcgtgcg gctccgcgct gcccggcggc tgtgagcgct gcgggcgcgg
     2941 cgcggggctt tgtgcgctcc gcagtgtgcg cgaggggagc gcggccgggg gcggtgcccc
     3001 gcggtgcggg gggggctgcg aggggaacaa aggctgcgtg cggggtgtgt gcgtgggggg
     3061 gtgagcaggg ggtgtgggcg cgtcggtcgg gctgcaaccc cccctgcacc cccctccccg
     3121 agttgctgag cacggcccgg cttcgggtgc ggggctccgt acggggcgtg gcgcggggct
     3181 cgccgtgccg ggcggggggt ggcggcaggt gggggtgccg ggcggggcgg ggccgcctcg
     3241 ggccggggag ggctcggggg aggggcgcgg cggcccccgg agcgccggcg gctgtcgagg
     3301 cgcggcgagc cgcagccatt gccttttatg gtaatcgtgc gagagggcgc agggacttcc
     3361 tttgtcccaa atctgtgcgg agccgaaatc tgggaggcgc cgccgcaccc cctctagcgg
     3421 gcgcggggcg aagcggtgcg gcgccggcag gaaggaaatg ggcggggagg gccttcgtgc
     3481 gtcgccgcgc cgccgtcccc ttctccctct ccagcctcgg ggctgtccgc ggggggacgg
     3541 ctgccttcgg gggggacggg gcagggcggg gttcggcttc tggcgtgtga ccggcggctc
     3601 tagagcctct gctaaccatg ttcatgcctt cttctttttc ctacagctcc tgggcaacgt
     3661 gctggttatt gtgctgtctc atcattttgg caaagaattg ctcgatcgac ggtatcgatg
     3721 ccaccatgac ggcgtctccc agtgggaagg gagcccgggc gctggaggct cctggctgcg
     3781 ggccccggcc gctcgcccgg gacctggtgg actccgtgga cgatgcggag gggctgtacg
     3841 tggctgtgga gcgctgcccg ctgtgcaaca ctacccgccg gcggctgacc tgcgccaaat
     3901 gcgttcagag cggcgatttc gtctacttcg acggccgcga ccgggagagg tttatcgaca
     3961 agaaggaaag gttaagccga cttaagagca agcaagaaga atttcagaaa gaagtgttaa
     4021 aagctatgga aggaaaatgg ataacagatc agttgagatg gaaaataatg tcctgcaaga
     4081 tgaggattga acagttaaaa caaacaatat gtaaaggaaa tgaagaaatg gagaaaaatt
     4141 ctgaaggcct tctcaaaacc aaggaaaaga atcagaagct ttacagtcga gcacaacggc
     4201 accaagagaa aaaggagaag attcagaggc ataatcgcaa acttggtgac ctggtagaaa
     4261 aaaagaccat tgacttaaga agtcattatg agcgtctggc aaatcttcga cgatcccata
     4321 tattagagct cacctctgtc atttttccaa tcgaggaagt aaagacgggt gtgagagacc
     4381 ccgcagatgt gtcttcagag agtgacagtg ccatgacctc cagcactgtg agcaagcttg
     4441 ctgaagcccg gaggacaact tacctctcag gacgatgggt ctgtgacgat cacagcggag
     4501 acaccagcat tagcattaca gggccttgga ttagcctccc taacaatggg gactactctg
     4561 cctactacag ctgggtggag gagaagaaaa caacccaggg gcctgacatg gagcagagta
     4621 accctgccta caccatcagt gctgcgctgt gctatgcaac tcagctggtc aacattctgt
     4681 ctcatatact tgatgtaaat cttcccaaaa agctctgcaa cagtgaattt tgtggcgaaa
     4741 atctaagcaa gcagaaattt actcgagcag tgaagaaact gaatgcaaat attctttacc
     4801 tttgtttttc tcagcatgta aatttagatc aattacaacc actgcatacc ctcaggaatc
     4861 taatgtacct ggtcagtcca agctctgaac acctaggcag gtcagggccc tttgaagtac
     4921 gagcagacct tgaggagtcc atggaatttg tggatcccgg agttgctgga gaatcagatg
     4981 agagcggaga tgagcgcgtc agcgatgaag aaaccgacct gggcacagac tgggagaact
     5041 tgcctagtcc ccggttttgt gatatccctt cccagtctgt ggaagtctcc cagagtcaga
     5101 gcacccaggc gtccccaccc atcgcgagca gcagtgcagg tgggatgatc tcctctgcag
     5161 cagcctcggt gacctcctgg tttaaagctt acactggaca ccgttaactc gaggaattca
     5221 ctcctcaggt gcaggctgcc tatcagaagg tggtggctgg tgtggccaat gccctggctc
     5281 acaaatacca ctgagatctt tttccctctg ccaaaaatta tggggacatc atgaagcccc
     5341 ttgagcatct gacttctggc taataaagga aatttatttt cattgcaata gtgtgttgga
     5401 attttttgtg tctctcactc ggaaggacat atgggagggc aaatcattta aaacatcaga
     5461 atgagtattt ggtttagagt ttggcaacat atgcccatat gctggctgcc atgaacaaag
     5521 gttggctata aagaggtcat cagtatatga aacagccccc tgctgtccat tccttattcc
     5581 atagaaaagc cttgacttga ggttagattt tttttatatt ttgttttgtg ttattttttt
     5641 ctttaacatc cctaaaattt tccttacatg ttttactagc cagatttttc ctcctctcct
     5701 gactactccc agtcatagct gtccctcttc tcttatggag atccctcgac ctgcagccca
     5761 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     5821 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     5881 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     5941 cagcggatcc gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     6001 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     6061 tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct
     6121 tttttggagg cctaggcttt tgcaaaaagc taacttgttt attgcagctt ataatggtta
     6181 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     6241 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggatccgct gcattaatga
     6301 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     6361 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     6421 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     6481 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     6541 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     6601 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     6661 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     6721 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     6781 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     6841 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     6901 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     6961 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     7021 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     7081 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     7141 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     7201 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     7261 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     7321 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     7381 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     7441 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     7501 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     7561 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     7621 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     7681 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     7741 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     7801 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     7861 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     7921 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     7981 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     8041 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     8101 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     8161 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     8221 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctg
//
