LOCUS       sequence-342320-        8730 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Vector for expression of 3xFLAG-tagged wild-type AMBRA1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8730)
  AUTHORS   Chaikovsky AC, Li C, Jeng EE, Loebell S, Lee MC, Murray CW, Cheng R,
            Demeter J, Swaney DL, Chen SH, Newton BW, Johnson JR, Drainas AP, 
            Shue YT, Seoane JA, Srinivasan P, He A, Yoshida A, Hipkins SQ, 
            McCrea E, Poltorack CD, Krogan NJ, Diehl JA, Kong C, Jackson PK, 
            Curtis C, Petrov DA, Bassik MC, Winslow MM, Sage J
  TITLE     The AMBRA1 E3 ligase adaptor regulates the stability of cyclin D.
  JOURNAL   Nature. 2021 Apr;592(7856):794-798. doi: 10.1038/s41586-021-03474-7.
            Epub 2021 Apr 14.
  PUBMED    33854239
REFERENCE   2  (bases 1 to 8730)
  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"; doi: 
            "10.1038/s41586-021-03474-7"; journalName: "Nature"; date: 
            "2021-04"; volume: "592"; issue: "7856"; pages: "794-798"
COMMENT     Sequence Label: pcDNA4-AMBRA1-3xFLAG
FEATURES             Location/Qualifiers
     source          1..8730
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    820..838
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    841..859
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     869..893
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             4632..4697
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(4738..4755)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4744..4968
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      5014..5442
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5101..5120)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5311..5332
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5451..5471)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5456..5785
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5636..5771
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5698..5717
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        5833..5880
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5899..6273
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    6403..6524
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6440..6459)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6494..6513
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6573..6589)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6573..6589)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6586..6608)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6597..6613
                     /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(6621..6627,6628..6645,6646..6651))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6666..6687
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6804..6821)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6975..7563)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7055..7074)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7734..8525,8526..8594))
                     /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     8357..8376
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8595..8699)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc
      841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga
      901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga
      961 ctctagcgtt taaacttaag cttgatatcg ccaccatgaa ggttgtccca gaaaagaatg
     1021 ctgtccggat actctggggg cgagaacggg gtgctcgggc catgggagct cagcggcttc
     1081 tgcaggagct ggtagaggat aaaacccggt ggatgaaatg ggagggcaag agagtagaac
     1141 tgccggatag tccacgctct accttcttat tggccttcag cccagacagg actctcttag
     1201 cctccaccca tgtgaaccat aatatctata ttacggaggt gaagactggc aagtgtgttc
     1261 attccctgat tggacaccgc cgtactccat ggtgtgtcac ttttcatccc accatctcag
     1321 gccttattgc ttctggctgc ctagatgggg aggttaggat ttgggattta cacggtggca
     1381 gtgaaagctg gttcacagat agcaacaatg ccattgcctc cctggctttc caccctacgg
     1441 ctcagctcct gctgattgcc actgccaatg agatccactt ctgggactgg agtcgacggg
     1501 aaccctttgc tgtggtgaag acagctagtg agatggaacg ggtccgtctg gtgagatttg
     1561 atccacttgg acactactta ctcacagcaa ttgttaaccc ctctaatcaa cagggtgatg
     1621 acgaaccaga gatccccata gatggaacag aattatccca ctaccgtcag cgtgccctcc
     1681 tgcaatcaca gccagttcgc cggacgcctc tcctccacaa tttcctgcac atgctgtcct
     1741 cccgctcttc tggcatccag accgagccct tccatccccc ggagcaggcc tcgtcaacgc
     1801 agcaggacca gggcctcctg aaccggccgt ctgccttcag tacagtccag agcagcactg
     1861 ccggcaacac gctccgcaac ctcagtctgg gtcctacccg ccgctctttg ggagggcctc
     1921 tgtctagcca cccttctagg tatcaccgag aaatagctcc tgggttgaca ggatctgagt
     1981 ggacccggac agtactcagt ctgaactccc gctctgaggc ggaatccatg cccccgccca
     2041 gaaccagtgc ctcttcggtg agtttgctgt ctgtgctgag acagcaggaa ggtggctctc
     2101 aggcatctgt gtacacttca gccacagaag ggaggggttt tccggcatca gggttggcaa
     2161 ctgagtcaga tggagggaat ggctccagcc aaaacaactc gggcagcatt cgccatgagc
     2221 ttcagtgtga cctgagacgc ttctttctgg agtatgaccg gcttcaggag ctggatcaga
     2281 gcctgagtgg ggaagctccc cagacccaac aggcccagga aatgctcaac aataacattg
     2341 aatctgagag gccaggccct tcccaccagc ccaccccaca cagcagtgag aacaactcca
     2401 acctgtcccg tggccacctg aatcgctgtc gtgcttgcca caatctcctg accttcaaca
     2461 acgataccct gcgctgggaa agaaccacac ctaactactc ctctggcgag gctagttcct
     2521 cttggcaggt ccccagctcc tttgagagtg tgccatcaag tggcagccag ttgccacctc
     2581 tcgagcggac tgagggccaa acgcccagct ccagcaggct ggagttgagc agctctgcta
     2641 gtccgcagga ggagaggact gtgggggtgg cctttaacca ggagacaggc cactgggaaa
     2701 gaatttacac ccagtccagc agatctggaa ctgtgtcaca ggaggcctta catcaggata
     2761 tgcctgagga gagctctgag gaggattcac tcaggaggag gctgctggaa tcttccctca
     2821 tttcattatc ccgttatgat ggagcaggat ccagagagca cccaatttac ccagacccag
     2881 cgagattatc tcctgctgca tactacgccc agaggatgat ccagtatctc tcacggagag
     2941 acagtattcg ccagcgctcc atgcgctacc aacagaaccg tctccgttct tccacctcct
     3001 cctcttcctc agacaaccag ggtccatcag tagagggaac cgacttggaa tttgaggact
     3061 ttgaggacaa tggtgacaga tccaggcacc gagctccacg caatgcccgg atgtctgcac
     3121 cttcgcttgg acgctttgtc ccaaggcgtt tcttgctgcc tgagtacttg ccttatgctg
     3181 ggatttttca tgaacgtgga cagcctggct tggctactca ctcttctgtt aacagggtcc
     3241 tggcaggggc agtgatcggt gatggacagt ctgctgtggc cagtaacatt gccaatacta
     3301 cctaccggct ccagtggtgg gacttcacta agtttgacct ccctgaaatc agtaatgctt
     3361 ccgtgaatgt gctggtgcag aactgcaaga tctacaatga tgccagctgt gacatttctg
     3421 cagatggcca gctcctggca gctttcatcc ccagcagcca gaggggcttt cctgatgaag
     3481 gcatcctggc agtgtactcc ctggcccccc ataacctggg cgaaatgctc tacaccaagc
     3541 gatttggtcc caatgccatt tcggtgagcc tgtccccaat gggcagatat gtaatggtgg
     3601 gcttggcctc acgaaggatc ctgctgcacc cctccacaga gcacatggtg gcccaggtct
     3661 tcaggctgca acaggcccat ggtggagaga cctccatgag gagagttttc aacgtccttt
     3721 atcccatgcc tgccgaccag cggagacatg tcagtatcaa ctctgcccgt tggctgcctg
     3781 agccagggct tggcttggcc tatggtacta acaaaggaga cctggtgatc tgccgaccag
     3841 aggccttaaa ctctggtgtt gagtactact gggaccagct gaacgagacg gtcttcactg
     3901 tccattccaa cagcaggagc agcgagcggc ctggaaccag cagagccaca tggaggacag
     3961 acagagacat ggggctgatg aatgccattg ggcttcagcc ccggaaccct gccacctcag
     4021 tgacatctca gggcaccgag actctggccc ttcagctgca gaatgccgaa acacagactg
     4081 agagggaggt gccggagcca gggacagccg cctcaggtcc tggtgaaggt gagggttcag
     4141 agtatggtgc cagtggagaa gatgcgctca gcaggatcca gaggctgatg gcggagggcg
     4201 gcatgacagc cgtggtgcag cgggagcaga gcaccaccat ggcctccatg ggcggcttcg
     4261 gcaacaacat catcgtcagc caccgcattc accgcagctc tcagacgggc actgaacctg
     4321 gtgccgccca cacctcctca ccccagccct ccacctctcg gggactgctc ccagaggccg
     4381 ggcaactggc agagcgaggc ctaagccccc ggacagcttc ctgggaccag cctggtaccc
     4441 ctgggcggga gccaacccag ccaaccctgc cctcttcctc ccctgtcccc attcctgttt
     4501 cccttcccag cgctgaggga ccaaccgtcc actgcgagtt gaccaataac aaccaccttc
     4561 tggatggtgg cagcagcagg ggggacgctg caggccctag gggagaacca cggaacaggg
     4621 gcggccgcat ggactacaag gaccacgacg gcgactacaa ggaccacgac atcgactaca
     4681 aggacgacga cgacaagtaa ctcgagtcta gagggcccgt ttaaacccgc tgatcagcct
     4741 cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga
     4801 ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt
     4861 gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg
     4921 attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg
     4981 aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg
     5041 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     5101 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     5161 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     5221 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     5281 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     5341 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     5401 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg
     5461 tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     5521 tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     5581 gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc
     5641 gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat
     5701 ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt
     5761 ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc
     5821 tgatcagcac gtgttgacaa ttaatcatcg gcatagtata tcggcatagt ataatacgac
     5881 aaggtgagga actaaaccat ggccaagttg accagtgccg ttccggtgct caccgcgcgc
     5941 gacgtcgccg gagcggtcga gttctggacc gaccggctcg ggttctcccg ggacttcgtg
     6001 gaggacgact tcgccggtgt ggtccgggac gacgtgaccc tgttcatcag cgcggtccag
     6061 gaccaggtgg tgccggacaa caccctggcc tgggtgtggg tgcgcggcct ggacgagctg
     6121 tacgccgagt ggtcggaggt cgtgtccacg aacttccggg acgcctccgg gccggccatg
     6181 accgagatcg gcgagcagcc gtgggggcgg gagttcgccc tgcgcgaccc ggccggcaac
     6241 tgcgtgcact tcgtggccga ggagcaggac tgacacgtgc tacgagattt cgattccacc
     6301 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
     6361 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct
     6421 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     6481 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg
     6541 tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt
     6601 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
     6661 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
     6721 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     6781 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     6841 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     6901 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     6961 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     7021 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     7081 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     7141 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     7201 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     7261 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     7321 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     7381 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg
     7441 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     7501 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     7561 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     7621 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     7681 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     7741 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     7801 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     7861 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     7921 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     7981 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     8041 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     8101 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     8161 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     8221 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     8281 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     8341 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     8401 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     8461 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     8521 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     8581 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     8641 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     8701 acatttcccc gaaaagtgcc acctgacgtc
//
