LOCUS       sequence-209822-        7756 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  donor vector for knock-in gene editing at human DNM2 locus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7756)
  AUTHORS   Srinivasan S, Burckhardt CJ, Bhave M, Chen Z, Chen PH, Wang X, 
            Danuser G, Schmid SL
  TITLE     A noncanonical role for dynamin-1 in regulating early stages of 
            clathrin-mediated endocytosis in non-neuronal cells.
  JOURNAL   PLoS Biol. 2018 Apr 18;16(4):e2005377. doi: 
            10.1371/journal.pbio.2005377. eCollection 2018 Apr.
  PUBMED    29668686
REFERENCE   2  (bases 1 to 7756)
  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: "PLoS 
            Biol."; date: "2018-04-18"; volume: "16(4):e2005377. doi"; pages: " 
            10.1371/journal.pbio.2005377. eCollection 2018 Apr"
COMMENT     Sequence Label: pRS DV DNM2-mRUBY2
FEATURES             Location/Qualifiers
     source          1..7756
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     27..46
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        27..45
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             812..1519
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="VSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTMR
                     IKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGGV
                     VTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTHM
                     ALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAVA
                     KFAGLGGGMDELYK"
     primer_bind     complement(2309..2329)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2309..2327)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2348..2364)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2348..2364)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2361..2383)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2372..2388
                     /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(2396..2402,2403..2420,2421..2426))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2441..2462
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2579..2596)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2750..3338)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2830..2849)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3509..4300,4301..4369))
                     /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     4132..4151
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4370..4474)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4501..5843
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(5464..5511)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     primer_bind     5884..5902
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5940..5962)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6062..6081
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        6102..6383
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     CDS             6384..7058
                     /codon_start=1
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /label=TRP1
                     /note="yeast auxotrophic marker"
                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
                     KK"
     rep_origin      complement(7161..7616)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7298..7319)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7510..7529
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     join(7742..7756,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(7756,1..17)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
ORIGIN
        1 gtaaaacgac ggccagtgag cgcgcgtaat acgactcact atagggcgaa ttgggtaccg
       61 ccccgcctca gatcccatct cggccagttc ggatcccccc agggattccc ccaggagtgc
      121 ccaggtaagg ccaaccccct gccctccacc ccaactgcct gcaccctggg gtctctcctc
      181 ctgtctcact tcctcccagt gagctctcac tacgtgccca gctgctggag tggaggaatc
      241 gtcctcatcc ctatttggct tgcgaggaaa caggcccaga gaggccaaga ggcttgttca
      301 gtgtcacaga gtagcggagc cctggtgact ccggggctcc cagcttgccc tctgcctact
      361 ggggtgtgcc accaggcagc tggggaaccc tcacactggg cacctcctcc cactgtttac
      421 cttcttctcc ttcctgctcc tgcctgcctc aacctaccgt gggccaggct ttattctccc
      481 cacttctccg gttgggaagc tgaggcccag ggagggcagg gggctcactg agggtcaaac
      541 agcaggcgag ttgatgccta ggtttggcac tcctgcccga caccctggtg gcttgcggag
      601 gtcagcctgg taacaccctg gggtggtgtg tgggtggggg ctgggtcctc aacccaggcc
      661 tgcctcttgc tcccggcctc actgccgtct ccccctcccc acctgtcttt attctctttg
      721 cagcagaaga ccccctgctg cgcccagccg gcccaccatt atccgcccag ccgagccatc
      781 cctgctcgac gacccgccgg tggcgacgtt ggtgagcaag ggcgaagagc tgatcaagga
      841 aaatatgcgt atgaaggtgg tcatggaagg ttcggtcaac ggccaccaat tcaaatgcac
      901 aggtgaagga gaaggcaatc cgtacatggg aactcaaacc atgaggatca aagtcatcga
      961 gggaggaccc ctgccatttg cctttgacat tcttgccacg tcgttcatgt atggcagccg
     1021 tacttttatc aagtacccga aaggcattcc tgatttcttt aaacagtcct ttcctgaggg
     1081 ttttacttgg gaaagagtta cgagatacga agatggtgga gtcgtcaccg tcatgcagga
     1141 caccagcctt gaggatggct gtctcgttta ccacgtccaa gtcagagggg taaactttcc
     1201 ctccaatggt cccgtgatgc agaagaagac caagggttgg gagcctaata cagagatgat
     1261 gtatccagca gatggtggtc tgaggggata cactcatatg gcactgaaag ttgatggtgg
     1321 tggccatctg tcttgctctt tcgtaacaac ttacaggtca aaaaagaccg tcgggaacat
     1381 caagatgccc ggtatccatg ccgttgatca ccgcctggaa aggttagagg aaagtgacaa
     1441 tgaaatgttc gtagtacaac gcgaacacgc agttgccaag ttcgccgggc ttggtggtgg
     1501 gatggacgag ctgtacaagt aagcctcgag gggggcgtgc tctcgggggg gcctcacgta
     1561 ttcgcggcac tagtgcttca gtggtctggg gccctccgcc gcccctatgc tgggaccagg
     1621 ctcccagtgg gcagccctgg cctcttcctt aacgctggcc ccggtccagg gccggcccct
     1681 gtgcctggct ggacaccgca ctgcgcaaag gggccctgga gctccaggca gggggcgctg
     1741 gggtgttgca ctttggggga tggagtctca gggtggcaga ggggggacca gaacccttga
     1801 caccatcctg aatgaggggt ccagcctggg ggggactcta ccaaggtctt cttgggctgg
     1861 gaaagcccat gtagggcagg ccttctataa gtgcgggcac caagggcgcc tacatcccca
     1921 ggccttgctg gggtgcaggg gtatatcaac ttcccattag caggagctcc ccagcggcaa
     1981 gcctggccca gtgggctcgg tagtgcccag ctggcaggcc tgaggtgtac atagtccttc
     2041 ccggccatat taaccacaca gcctgagcct ggcccagcct cggctgccag aggtgccttt
     2101 gctaggcccg gagccgttgg cccgggccgg ccttgcccta ttcctctcct cctcctcctc
     2161 ctgggtcccc cagggtggct gggcttgggc tatgtgggtg gtggtggcgg ggggtcttgg
     2221 gggcctctca gctcccgccc atgcctccct gatgggtggg cccagggcgg ccgcggccgc
     2281 caccgcggtg gagctccagc ttttgttccc tttagtgagg gttaattgcg cgcttggcgt
     2341 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     2401 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     2461 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     2521 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     2581 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     2641 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     2701 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     2761 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     2821 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     2881 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     2941 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     3001 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     3061 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     3121 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     3181 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     3241 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     3301 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     3361 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     3421 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     3481 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     3541 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     3601 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     3661 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     3721 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     3781 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     3841 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     3901 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     3961 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     4021 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     4081 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     4141 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     4201 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     4261 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     4321 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     4381 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     4441 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     4501 aacgaagcat ctgtgcttca ttttgtagaa caaaaatgca acgcgagagc gctaattttt
     4561 caaacaaaga atctgagctg catttttaca gaacagaaat gcaacgcgaa agcgctattt
     4621 taccaacgaa gaatctgtgc ttcatttttg taaaacaaaa atgcaacgcg agagcgctaa
     4681 tttttcaaac aaagaatctg agctgcattt ttacagaaca gaaatgcaac gcgagagcgc
     4741 tattttacca acaaagaatc tatacttctt ttttgttcta caaaaatgca tcccgagagc
     4801 gctatttttc taacaaagca tcttagatta ctttttttct cctttgtgcg ctctataatg
     4861 cagtctcttg ataacttttt gcactgtagg tccgttaagg ttagaagaag gctactttgg
     4921 tgtctatttt ctcttccata aaaaaagcct gactccactt cccgcgttta ctgattacta
     4981 gcgaagctgc gggtgcattt tttcaagata aaggcatccc cgattatatt ctataccgat
     5041 gtggattgcg catactttgt gaacagaaag tgatagcgtt gatgattctt cattggtcag
     5101 aaaattatga acggtttctt ctattttgtc tctatatact acgtatagga aatgtttaca
     5161 ttttcgtatt gttttcgatt cactctatga atagttctta ctacaatttt tttgtctaaa
     5221 gagtaatact agagataaac ataaaaaatg tagaggtcga gtttagatgc aagttcaagg
     5281 agcgaaaggt ggatgggtag gttatatagg gatatagcac agagatatat agcaaagaga
     5341 tacttttgag caatgtttgt ggaagcggta ttcgcaatat tttagtagct cgttacagtc
     5401 cggtgcgttt ttggtttttt gaaagtgcgt cttcagagcg cttttggttt tcaaaagcgc
     5461 tctgaagttc ctatactttc tagagaatag gaacttcgga ataggaactt caaagcgttt
     5521 ccgaaaacga gcgcttccga aaatgcaacg cgagctgcgc acatacagct cactgttcac
     5581 gtcgcaccta tatctgcgtg ttgcctgtat atatatatac atgagaagaa cggcatagtg
     5641 cgtgtttatg cttaaatgcg tacttatatg cgtctattta tgtaggatga aaggtagtct
     5701 agtacctcct gtgatattat cccattccat gcggggtatc gtatgcttcc ttcagcacta
     5761 ccctttagct gttctatatg ctgccactcc tcaattggat tagtctcatc cttcaatgct
     5821 atcatttcct ttgatattgg atcatactaa gaaaccatta ttatcatgac attaacctat
     5881 aaaaataggc gtatcacgag gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac
     5941 ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc
     6001 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg gcttaactat
     6061 gcggcatcag agcagattgt actgagagtg caccatagat caacgacatt actatatata
     6121 taatatagga agcatttaat agaacagcat cgtaatatat gtgtactttg cagttatgac
     6181 gccagatggc agtagtggaa gatattcttt attgaaaaat agcttgtcac cttacgtaca
     6241 atcttgatcc ggagcttttc tttttttgcc gattaagaat taattcggtc gaaaaaagaa
     6301 aaggagaggg ccaagaggga gggcattggt gactattgag cacgtgagta tacgtgatta
     6361 agcacacaaa ggcagcttgg agtatgtctg ttattaattt cacaggtagt tctggtccat
     6421 tggtgaaagt ttgcggcttg cagagcacag aggccgcaga atgtgctcta gattccgatg
     6481 ctgacttgct gggtattata tgtgtgccca atagaaagag aacaattgac ccggttattg
     6541 caaggaaaat ttcaagtctt gtaaaagcat ataaaaatag ttcaggcact ccgaaatact
     6601 tggttggcgt gtttcgtaat caacctaagg aggatgtttt ggctctggtc aatgattacg
     6661 gcattgatat cgtccaactg catggagatg agtcgtggca agaataccaa gagttcctcg
     6721 gtttgccagt tattaaaaga ctcgtatttc caaaagactg caacatacta ctcagtgcag
     6781 cttcacagaa acctcattcg tttattccct tgtttgattc agaagcaggt gggacaggtg
     6841 aacttttgga ttggaactcg atttctgact gggttggaag gcaagagagc cccgaaagct
     6901 tacattttat gttagctggt ggactgacgc cagaaaatgt tggtgatgcg cttagattaa
     6961 atggcgttat tggtgttgat gtaagcggag gtgtggagac aaatggtgta aaagactcta
     7021 acaaaatagc aaatttcgtc aaaaatgcta agaaataggt tattactgag tagtatttat
     7081 ttaagtattg tttgtgcact tgccgatcta tgcggtgtga aataccgcac agatgcgtaa
     7141 ggagaaaata ccgcatcagg aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa
     7201 tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa
     7261 atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact
     7321 attaaagaac gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc
     7381 actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa
     7441 tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc
     7501 gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt
     7561 cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccat
     7621 tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta
     7681 cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac gccagggttt
     7741 tcccagtcac gacgtt
//
