LOCUS       sequence-272122-        6335 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the Beclin-1 IDR+CC domains (a.a. 1-265) with a N-terminal
            hexa-Histidine tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6335)
  AUTHORS   Ranaghan MJ, Durney MA, Mesleh MF, McCarren PR, Garvie CW, Daniels 
            DS, Carey KL, Skepner AP, Levine B, Perez JR
  TITLE     The Autophagy-Related Beclin-1 Protein Requires the Coiled-Coil and 
            BARA Domains To Form a Homodimer with Submicromolar Affinity.
  JOURNAL   Biochemistry. 2017 Dec 26;56(51):6639-6651. doi: 
            10.1021/acs.biochem.7b00936. Epub 2017 Dec 14.
  PUBMED    29185708
REFERENCE   2  (bases 1 to 6335)
  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: 
            "10.1021/acs.biochem.7b00936"; journalName: "Biochemistry"; date: 
            "2017-12-26- 26"; volume: "56"; issue: "51"; pages: "6639-6651"
COMMENT     Sequence Label: BPV00605 pET21GG2-His(6)-BECN(1-265)
FEATURES             Location/Qualifiers
     source          1..6335
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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)."
     RBS             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             95..112
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             119..136
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             143..163
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             983..1006
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1007..1051
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             1067..1084
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1137..1155)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1151..1198
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1235..1690
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1322..1341)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1532..1553
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1716..1820
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1821..1889,1890..2681)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2039..2058)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2852..3440
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3341..3360
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3594..3611
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3626..3765
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3691..3710)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3851..3873
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3867..4058)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4867..5949)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5911..5930
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5950..6027)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6233..6252
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat gagccatcat catcaccatc atggatccct
      121 ggttccgcgt ggtagcgcta gcgaaaatct gtattttcag ggaatggaag gtagcaaaac
      181 cagcaataat agcaccatgc aggttagctt tgtttgtcag cgttgtagcc agccgctgaa
      241 actggatacc agctttaaaa tcctggatcg tgttaccatt caagaactga ccgcaccgct
      301 gctgaccacc gcacaggcaa aaccgggtga aacccaagaa gaagaaacca attcaggcga
      361 agaaccgttt attgaaacac cgcgtcagga tggtgttagc cgtcgtttta ttccgcctgc
      421 acgtatgatg agcaccgaaa gcgcaaatag ctttaccctg attggtgaag caagtgatgg
      481 tggcaccatg gaaaatctga gccgtcgtct gaaagttacc ggtgacctgt ttgatatcat
      541 gagcggtcag accgatgttg atcatccgct gtgtgaagaa tgtaccgata ccctgctgga
      601 tcagctggat acccaactga atgttaccga aaatgaatgc cagaactata aacgctgtct
      661 ggaaattctg gaacaaatga acgaagatga tagcgaacag ctgcagatgg aactgaaaga
      721 actggcactg gaagaagaac gtctgatcca agaactggaa gatgtggaaa aaaaccgtaa
      781 aattgtggcc gagaatctgg aaaaagttca ggcagaagcc gaacgcctgg atcaagaaga
      841 ggcacagtat cagcgtgaat atagcgaatt taaacgtcag cagctggaac tggacgatga
      901 actgaaaagc gttgaaaatc agatgcgtta tgcacagacc cagctggaca aactgaaata
      961 aagcctcgag ggtggtggta gcgattataa agatgatgat gataaaggcc tgaacgatat
     1021 ctttgaagcc cagaaaattg aatggcacga gtaaaagctt gtcgagcacc accaccacca
     1081 ccactgagat ccggctgcta acaaagcccg aaaggaagct gagttggctg ctgccaccgc
     1141 tgagcaataa ctagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgct
     1201 gaaaggagga actatatccg gattggcgaa tgggacgcgc cctgtagcgg cgcattaagc
     1261 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     1321 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     1381 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     1441 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     1501 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     1561 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     1621 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     1681 cttacaattt aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     1741 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     1801 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     1861 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     1921 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     1981 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     2041 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     2101 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     2161 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     2221 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     2281 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     2341 acgagcgtga caccacgatg cctgcagcaa tggcaacaac gttgcgcaaa ctattaactg
     2401 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     2461 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     2521 gagccggtga gcgtggtagt cgcggtatca ttgcagcact ggggccagat ggtaagccct
     2581 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     2641 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     2701 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     2761 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     2821 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     2881 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     2941 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     3001 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     3061 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     3121 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     3181 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     3241 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     3301 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     3361 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     3421 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     3481 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     3541 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     3601 cagtgagcga ggaagcggaa gagcgcctga tgcggtattt tctccttacg catctgtgcg
     3661 gtatttcaca ccgcaatggt gcactctcag tacaatctgc tctgatgccg catagttaag
     3721 ccagtataca ctccgctatc gctacgtgac tgggtcatgg ctgcgccccg acacccgcca
     3781 acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct
     3841 gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg
     3901 aggcagctgc ggtaaagctc atcagcgtgg tcgtgaagcg attcacagat gtctgcctgt
     3961 tcatccgcgt ccagctcgtt gagtttctcc agaagcgtta atgtctggct tctgataaag
     4021 cgggccatgt taagggcggt tttttcctgt ttggtcactg atgcctccgt gtaaggggga
     4081 tttctgttca tgggggtaat gataccgatg aaacgagaga ggatgctcac gatacgggtt
     4141 actgatgatg aacatgcccg gttactggaa cgttgtgagg gtaaacaact ggcggtatgg
     4201 atgcggcggg accagagaaa aatcactcag ggtcaatgcc agcgcttcgt taatacagat
     4261 gtaggtgttc cacagggtag ccagcagcat cctgcgatgc agatccggaa cataatggtg
     4321 cagggcgctg acttccgcgt ttccagactt tacgaaacac ggaaaccgaa gaccattcat
     4381 gttgttgctc aggtcgcaga cgttttgcag cagcagtcgc ttcacgttcg ctcgcgtatc
     4441 ggtgattcat tctgctaacc agtaaggcaa ccccgccagc ctagccgggt cctcaacgac
     4501 aggagcacga tcatgcgcac ccgtggggcc gccatgccgg cgataatggc ctgcttctcg
     4561 ccgaaacgtt tggtggcggg accagtgacg aaggcttgag cgagggcgtg caagattccg
     4621 aataccgcaa gcgacaggcc gatcatcgtc gcgctccagc gaaagcggtc ctcgccgaaa
     4681 atgacccaga gcgctgccgg cacctgtcct acgagttgca tgataaagaa gacagtcata
     4741 agtgcggcga cgatagtcat gccccgcgcc caccggaagg agctgactgg gttgaaggct
     4801 ctcaagggca tcggtcgaga tcccggtgcc taatgagtga gctaacttac attaattgcg
     4861 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     4921 ggccaacgcg cggggagagg cggtttgcgt attgggcgcc agggtggttt ttcttttcac
     4981 cagtgagacg ggcaacagct gattgccctt caccgcctgg ccctgagaga gttgcagcaa
     5041 gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg
     5101 gatataacat gagctgtctt cggtatcgtc gtatcccact accgagatat ccgcaccaac
     5161 gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc gccatctgat cgttggcaac
     5221 cagcatcgca gtgggaacga tgccctcatt cagcatttgc atggtttgtt gaaaaccgga
     5281 catggcactc cagtcgcctt cccgttccgc tatcggctga atttgattgc gagtgagata
     5341 tttatgccag ccagccagac gcagacgcgc cgagacagaa cttaatgggc ccgctaacag
     5401 cgcgatttgc tggtgaccca atgcgaccag atgctccacg cccagtcgcg taccgtcttc
     5461 atgggagaaa ataatactgt tgatgggtgt ctggtcagag acatcaagaa ataacgccgg
     5521 aacattagtg caggcagctt ccacagcaat ggcatcctgg tcatccagcg gatagttaat
     5581 gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc gccgctttac aggcttcgac
     5641 gccgcttcgt tctaccatcg acaccaccac gctggcaccc agttgatcgg cgcgagattt
     5701 aatcgccgcg acaatttgcg acggcgcgtg cagggccaga ctggaggtgg caacgccaat
     5761 cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg ttgggaatgt aattcagctc
     5821 cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa acgtggctgg cctggttcac
     5881 cacgcgggaa acggtctgat aagagacacc ggcatactct gcgacatcgt ataacgttac
     5941 tggtttcaca ttcaccaccc tgaattgact ctcttccggg cgctatcatg ccataccgcg
     6001 aaaggttttg cgccattcga tggtgtccgg gatctcgacg ctctccctta tgcgactcct
     6061 gcattaggaa gcagcccagt agtaggttga ggccgttgag caccgccgcc gcaaggaatg
     6121 gtgcatgcaa ggagatggcg cccaacagtc ccccggccac ggggcctgcc accataccca
     6181 cgccgaaaca agcgctcatg agcccgaagt ggcgagcccg atcttcccca tcggtgatgt
     6241 cggcgatata ggcgccagca accgcacctg tggcgccggt gatgccggcc acgatgcgtc
     6301 cggcgtagag gatcgagatc tcgatcccgc gaaat
//
