LOCUS       sequence-344446-        8612 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human Drp1 Isoform 3 with EGFP at N-terminus in bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8612)
  AUTHORS   Kamerkar SC, Kraus F, Sharpe AJ, Pucadyil TJ, Ryan MT
  TITLE     Dynamin-related protein 1 has membrane constricting and severing 
            abilities sufficient for mitochondrial and peroxisomal fission.
  JOURNAL   Nat Commun. 2018 Dec 7;9(1):5239. doi: 10.1038/s41467-018-07543-w.
  PUBMED    30531964
REFERENCE   2  (bases 1 to 8612)
  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: "Nat 
            Commun."; date: "2018-12-7"; volume: "9(1):5239. doi"; pages: " 
            10.1038/s41467-018-07543-w"
COMMENT     Sequence Label: 6xHis(Thrombin)-GFP(TEV)-Drp1-StrepII
FEATURES             Location/Qualifiers
     source          1..8612
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(1281..2363)
                     /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     2325..2344
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2364..2441)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     2647..2666
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     2750..2769
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2750..2768
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2769..2793
                     /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             2808..2830
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2849..2866
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2876..2893
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             join(2912..2914,2915..3625)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAEITLGMDELYK"
     primer_bind     complement(2954..2975)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3215..3234)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3562..3583
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3626..3646
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             5747..5770
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     primer_bind     complement(5862..5880)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      5876..5923
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(6098..6103,6104..6120,6121..6126))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(6153..6171)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6239..6343
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6344..6412,6413..7204)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(6562..6581)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7375..7963
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7864..7883
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8117..8134
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    8149..8288
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(8214..8233)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8374..8396
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(8390..8581)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 atgggggtaa tgataccgat gaaacgagag aggatgctca cgatacgggt tactgatgat
       61 gaacatgccc ggttactgga acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg
      121 gaccagagaa aaatcactca gggtcaatgc cagcgcttcg ttaatacaga tgtaggtgtt
      181 ccacagggta gccagcagca tcctgcgatg cagatccgga acataatggt gcagggcgct
      241 gacttccgcg tttccagact ttacgaaaca cggaaaccga agaccattca tgttgttgct
      301 caggtcgcag acgttttgca gcagcagtcg cttcacgttc gctcgcgtat cggtgattca
      361 ttctgctaac cagtaaggca accccgccag cctagccggg tcctcaacga caggagcacg
      421 atcatgcgca cccgtggcca ggacccaacg ctgcccgaga tgcgccgcgt gcggctgctg
      481 gagatggcgg acgcgatgga tatgttctgc caagggttgg tttgcgcatt cacagttctc
      541 cgcaagaatt gattggctcc aattcttgga gtggtgaatc cgttagcgag gtgccgccgg
      601 cttccattca ggtcgaggtg gcccggctcc atgcaccgcg acgcaacgcg gggaggcaga
      661 caaggtatag ggcggcgcct acaatccatg ccaacccgtt ccatgtgctc gccgaggcgg
      721 cataaatcgc cgtgacgatc agcggtccaa tgatcgaagt taggctggta agagccgcga
      781 gcgatccttg aagctgtccc tgatggtcgt catctacctg cctggacagc atggcctgca
      841 acgcgggcat cccgatgccg ccggaagcga gaagaatcat aatggggaag gccatccagc
      901 ctcgcgtcgc gaacgccagc aagacgtagc ccagcgcgtc ggccgccatg ccggcgataa
      961 tggcctgctt ctcgccgaaa cgtttggtgg cgggaccagt gacgaaggct tgagcgaggg
     1021 cgtgcaagat tccgaatacc gcaagcgaca ggccgatcat cgtcgcgctc cagcgaaagc
     1081 ggtcctcgcc gaaaatgacc cagagcgctg ccggcacctg tcctacgagt tgcatgataa
     1141 agaagacagt cataagtgcg gcgacgatag tcatgccccg cgcccaccgg aaggagctga
     1201 ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg tgcctaatga gtgagctaac
     1261 ttacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     1321 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgccagggtg
     1381 gtttttcttt tcaccagtga gacgggcaac agctgattgc ccttcaccgc ctggccctga
     1441 gagagttgca gcaagcggtc cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg
     1501 gtggttaacg gcgggatata acatgagctg tcttcggtat cgtcgtatcc cactaccgag
     1561 atatccgcac caacgcgcag cccggactcg gtaatggcgc gcattgcgcc cagcgccatc
     1621 tgatcgttgg caaccagcat cgcagtggga acgatgccct cattcagcat ttgcatggtt
     1681 tgttgaaaac cggacatggc actccagtcg ccttcccgtt ccgctatcgg ctgaatttga
     1741 ttgcgagtga gatatttatg ccagccagcc agacgcagac gcgccgagac agaacttaat
     1801 gggcccgcta acagcgcgat ttgctggtga cccaatgcga ccagatgctc cacgcccagt
     1861 cgcgtaccgt cttcatggga gaaaataata ctgttgatgg gtgtctggtc agagacatca
     1921 agaaataacg ccggaacatt agtgcaggca gcttccacag caatggcatc ctggtcatcc
     1981 agcggatagt taatgatcag cccactgacg cgttgcgcga gaagattgtg caccgccgct
     2041 ttacaggctt cgacgccgct tcgttctacc atcgacacca ccacgctggc acccagttga
     2101 tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg cgtgcagggc cagactggag
     2161 gtggcaacgc caatcagcaa cgactgtttg cccgccagtt gttgtgccac gcggttggga
     2221 atgtaattca gctccgccat cgccgcttcc actttttccc gcgttttcgc agaaacgtgg
     2281 ctggcctggt tcaccacgcg ggaaacggtc tgataagaga caccggcata ctctgcgaca
     2341 tcgtataacg ttactggttt cacattcacc accctgaatt gactctcttc cgggcgctat
     2401 catgccatac cgcgaaaggt tttgcgccat tcgatggtgt ccgggatctc gacgctctcc
     2461 cttatgcgac tcctgcatta ggaagcagcc cagtagtagg ttgaggccgt tgagcaccgc
     2521 cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac agtcccccgg ccacggggcc
     2581 tgccaccata cccacgccga aacaagcgct catgagcccg aagtggcgag cccgatcttc
     2641 cccatcggtg atgtcggcga tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc
     2701 ggccacgatg cgtccggcgt agaggatcga gatctcgatc ccgcgaaatt aatacgactc
     2761 actatagggg aattgtgagc ggataacaat tcccctctag aaataatttt gtttaacttt
     2821 aagaaggaga tataccatgg gcagcagcca tcatcatcat catcacagca gcggcctggt
     2881 gccgcgcggc agccatatgc tcgaggatcc ggtgagcaag ggcgaggagc tgttcaccgg
     2941 ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc
     3001 cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac
     3061 cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg
     3121 cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga
     3181 aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc
     3241 cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt
     3301 caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt
     3361 ctatatcatg gccgacaagc agaagaacgg catcaaggtg aacttcaaga tccgccacaa
     3421 catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga
     3481 cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccaagt tgagcaaaga
     3541 ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgagatcac
     3601 tctcggcatg gacgagctgt acaaggaaaa tctgtatttc caaggctcga tggaggcgct
     3661 aattcctgtc ataaacaagc tccaggacgt cttcaacacg gtgggcgccg acatcatcca
     3721 gctgcctcaa atcgtcgtag tgggaacgca gagcagcgga aagagctcag tgctagaaag
     3781 cctggtgggg agggacctgc ttcccagagg tactggaatt gtcacccgga gacctctcat
     3841 tctgcaactg gtccatgttt cacaagaaga taaacggaaa acaacaggag aagaaaatgg
     3901 ggtggaagca gaagaatggg gtaaatttct tcacaccaaa aataagcttt acacggattt
     3961 tgatgaaatt cgacaagaaa ttgaaaatga aacagaaaga atttcaggaa ataataaggg
     4021 agtaagccct gaaccaattc atcttaagat tttttcaccc aacgttgtca atttgacact
     4081 tgtggatttg ccaggaatga ccaaggtgcc tgtaggtgat caacctaagg atattgagct
     4141 tcaaatcaga gagctcattc ttcggttcat cagtaatcct aattccatta tcctcgctgt
     4201 cactgctgct aatacagata tggcaacatc agaggcactt aaaatttcaa gagaggtaga
     4261 tccagatggt cgcagaaccc tagctgtaat cactaaactt gatctcatgg atgcgggtac
     4321 tgatgccatg gatgtattga tgggaagggt tattccagtc aaacttggaa taattggagt
     4381 agttaacagg agccagctag atattaacaa caagaagagt gtaactgatt caatccgtga
     4441 tgagtatgct tttcttcaaa agaaatatcc atctctggcc aatagaaatg gaacaaagta
     4501 tcttgctagg actctaaaca ggttactgat gcatcacatc agagattgtt taccagagtt
     4561 gaaaacaaga ataaatgttc tagctgctca gtatcagtct cttctaaata gctacggtga
     4621 acccgtggat gataaaagtg ctactttact ccaacttatt accaaatttg ccacagaata
     4681 ttgtaacact attgaaggaa ctgcaaaata tattgaaact tcggagctat gcggtggtgc
     4741 tagaatttgt tatattttcc atgagacttt tgggcgaacc ttagaatctg ttgatccact
     4801 tggtggcctt aacactattg acattttgac tgccattaga aatgctactg gtcctcgtcc
     4861 tgctttattt gtgcctgagg tttcatttga gttactggtg aagcggcaaa tcaaacgtct
     4921 agaagagccc agcctccgct gtgtggaact ggttcatgag gaaatgcaaa ggatcattca
     4981 gcactgtagc aattacagta cacaggaatt gttacgattt cctaaacttc atgatgccat
     5041 agttgaagtg gtgacttgtc ttcttcgtaa aaggttgcct gttacaaatg aaatggtcca
     5101 taacttagtg gcaattgaac tggcttatat caacacaaaa catccagact ttgctgatgc
     5161 ttgtgggcta atgaacaata atatagagga acaaaggaga aacaggctag ccagagaatt
     5221 accttcagct gtatcacgag acaaggttgc atctggaggt ggtggggttg gagatggtgt
     5281 tcaagaacca accacaggca actggagagg aatgctgaaa acttcaaaag ctgaagagtt
     5341 attagcagaa gaaaaatcaa aacccattcc aattatgcca gccagtccac aaaaaggtca
     5401 tgccgtgaac ctgctagatg tgccagttcc tgttgcacga aaactatctg ctcgggaaca
     5461 gcgagattgt gaggttattg aacgactcat taaatcatat tttctcattg tcagaaagaa
     5521 tattcaagac agtgtgccaa aggcagtaat gcattttttg gttaatcatg tgaaagacac
     5581 tcttcagagt gagctagtag gccagctgta taaatcatcc ttattggatg atcttctgac
     5641 agaatctgag gacatggcac agcgcaggaa agaagcagct gatatgctaa aggcattaca
     5701 aggagccagt caaattattg ctgaaatccg ggagactcat ctttggtggt ctcatcctca
     5761 gttcgaaaaa tgaggcaggc cctcagccgc tttccagtga tgtagggtac cggatccggc
     5821 tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc accgctgagc aataactagc
     5881 ataacccctt ggggcctcta aacgggtctt gaggggtttt ttgctgaaag gaggaactat
     5941 atccggatat cccgcaagag gcccggcagt accggcataa ccaagcctat gcctacagca
     6001 tccagggtga cggtgccgag gatgacgatg agcgcattgt tagatttcat acacggtgcc
     6061 tgactgcgtt agcaatttaa ctgtgataaa ctaccgcatt aaagcttatc gatgataagc
     6121 tgtcaaacat gagaattctt gaagacgaaa gggcctcgtg atacgcctat ttttataggt
     6181 taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg
     6241 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     6301 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     6361 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     6421 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     6481 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     6541 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtgttg acgccgggca
     6601 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     6661 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     6721 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     6781 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     6841 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgcag caatggcaac
     6901 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     6961 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     7021 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     7081 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     7141 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     7201 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     7261 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     7321 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     7381 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     7441 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     7501 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa
     7561 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     7621 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     7681 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     7741 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     7801 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     7861 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     7921 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     7981 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     8041 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     8101 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta
     8161 ttttctcctt acgcatctgt gcggtatttc acaccgcaat ggtgcactct cagtacaatc
     8221 tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt gactgggtca
     8281 tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     8341 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     8401 caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgaa
     8461 gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc tccagaagcg
     8521 ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc tgtttggtca
     8581 ctgatgcctc cgtgtaaggg ggatttctgt tc
//
