LOCUS       sequence-373473-        5805 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Repair template for the N-terminal tagging of Ect2 with mNeonGreen 
            in human cells using CRISPR/Cas9..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5805)
  AUTHORS   Husser MC, Ozugergin I, Resta T, Martin VJJ, Piekny AJ
  TITLE     Cytokinetic diversity in mammalian cells is revealed by the 
            characterization of endogenous anillin, Ect2 and RhoA.
  JOURNAL   Open Biol. 2022 Nov;12(11):220247. doi: 10.1098/rsob.220247. Epub 
            2022 Nov 23.
  PUBMED    36416720
REFERENCE   2  (bases 1 to 5805)
  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: "Open 
            Biol."; date: "2022-11"; volume: "12(11):220247. doi"; pages: " 
            10.1098/rsob.220247. Epub 2022 Nov 23"
COMMENT     Sequence Label: pODN-mNG-ECT2
FEATURES             Location/Qualifiers
     source          1..5805
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(171..962,963..1031))
                     /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     794..813
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1032..1136)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1204..1222
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     1528..1547
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1528..1546
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2697..3404
                     /codon_start=1
                     /product="bright monomeric yellow-green fluorescent protein
                     derived from LanYFP (Shaner et al., 2013)"
                     /label=mNeonGreen
                     /translation="MVSKGEEDNMASLPATHELHIFGSINGVDFDMVGQGTGNPNDGYE
                     ELNLKSTKGDLQFSPWILVPHIGYGFHQYLPYPDGMSPFQAAMVDGSGYQVHRTMQFED
                     GASLTVNYRYTYEGSHIKGEAQVKGTGFPADGPVMTNSLTAADWCRSKKTYPNDKTIIS
                     TFKWSYTTGNGKRYRSTARTTYTFAKPMAANYLKNQPMYVFRKTELKHSKTELNFKEWQ
                     KAFTDVMGMDELYK"
     regulatory      2952..2961
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(join(4860..4866,4867..4884,4885..4890))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    4905..4926
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5046..5063)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5217..5805)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5297..5316)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
       61 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
      121 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
      181 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
      241 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
      301 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
      361 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
      421 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
      481 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
      541 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
      601 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
      661 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
      721 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
      781 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
      841 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
      901 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
      961 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     1021 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     1081 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     1141 tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac attaacctat
     1201 aaaaataggc gtatcacgag gccgcccctg cagccgaatt atattatttt tgccaaataa
     1261 tttttaacaa aagctctgaa gtcttcttca tttaaattct tagatgatac ttcatctgga
     1321 aaattgtccc aattagtagc atcacgctgt gagtaagttc taaaccattt ttttattgtt
     1381 gtattatctc taatcttact actcgatgag ttttcggtat tatctctatt tttaacttgg
     1441 agcaggttcc attcattgtt tttttcatca tagtgaataa aatcaactgc tttaacactt
     1501 gtgcctgaac accatatcca tccggcgtaa tacgactcac tatagggaga gcggccgcca
     1561 gatcttccgg atggctcgag tttttcagca agattctagc ttttaaactc ttcttaagtc
     1621 ctggaaaact taaaaaagga ttttctttca tagtccagag ttatattggc actcacccat
     1681 gcataattgt tctgtcaaac cttttagggg agaggtgttt ttaaagaatt atgaaaatac
     1741 tgacaaggga ccagcccaat tataccggag aggtttagag acatccacag aggatgagtc
     1801 aagagtgttg gattgggggt tggaagacaa gataggtgga gagtcgagca ctaaatgcaa
     1861 atagatgtga tgtaaattat gatttgtgcc tgaaaaatag ctgagaaatt cattgttgct
     1921 gttgcataga ttgtataggg actggggtag ttacagatgg tgagggtgaa tagttaggcg
     1981 gggaccaaat tgtgaacttg gtggtgaatt tggacttgga catataagga gtactcatgg
     2041 aagttgttaa gcagggaagt gatatggtca catttgtatt taggtgcata ttcctgaaaa
     2101 ccatgtggag agtgatttct aaggaaactg atggtaaagt tgtgtggagc tggtttatta
     2161 gaccaagtga ccattgtgtc attgaacgta tagagaaggg gggccagatt ccagagatac
     2221 ttgaggtggg attgatagga attttgggga gggaaatgtg ggagaggaaa aaattaggat
     2281 agatctcaga tttttggcta ggatttttgt gggaggtgtg ttatggtggt gttataattg
     2341 aatataggaa gagaaattgg ttgagggcaa aggtgtggag aagggaggta gaataatatt
     2401 tatgactaca gttgagtgtt tgttgtatgt ctagtaatgt tctatgtgct tttgatacat
     2461 tgttattttt attatattcc tgtgagtcag gtattagtat gtttatttta cagaaaaaga
     2521 aactgaggct cagagaaggt cagtgacttt ctttggttca ttaaaaaaat gggtaataat
     2581 actttttata aattctaggt aaaaagagcc tctaatactt acttttgccc aatgaccatg
     2641 tttatgtatt tttattcaaa ttttattttt tcagctgatt tagaagaata caaatcatgg
     2701 tgagcaaggg cgaggaggat aacatggcct ctctcccagc gacacatgag ttacacatct
     2761 ttggctccat caacggtgtg gactttgaca tggtgggtca gggcaccggc aatccaaatg
     2821 atggttatga ggagttaaac ctgaagtcca ccaagggtga cctccagttc tccccctgga
     2881 ttctggtccc tcatatcggg tatggcttcc atcagtacct gccctaccct gacgggatgt
     2941 cgcctttcca ggccgccatg gtagatggct ccggatacca agtccatcgc acaatgcagt
     3001 ttgaagatgg tgcctccctt actgttaact accgctacac ctacgaggga agccacatca
     3061 aaggagaggc ccaggtgaag gggactggtt tccctgctga cggtcctgtg atgaccaact
     3121 cgctgaccgc tgcggactgg tgcaggtcga agaagactta ccccaacgac aaaaccatca
     3181 tcagtacctt taagtggagt tacaccactg gaaatggcaa gcgctaccgg agcactgcgc
     3241 ggaccaccta cacctttgcc aagccaatgg cggctaacta tctgaagaac cagccgatgt
     3301 acgtgttccg taagacggag ctcaagcact ccaagaccga gctcaacttc aaggagtggc
     3361 aaaaggcctt taccgatgtg atgggcatgg acgagctgta caagggaggt tcaggaggct
     3421 ccatggcaga gaattcagta ctaacttcta cgacgggaag gactagcttg gcagactctt
     3481 ccatttttga ttctaaagtt actgagattt ccaaggaaaa cttacttatt ggatctactt
     3541 catatgtaga aggtaaacct gttacctgct ttaaaacaat caatttctat tttaattcta
     3601 aacttattag tgactttctt aagatttaat tttaatacca actgtaatgc tagagcacag
     3661 gtattaagta aatatttgtt tacagaatcc ttttacacta tgtcattttt ctgattatta
     3721 tttgggttat ttgtatgctt tgcagctttg aaaagggagg catcttaatt tggatattcc
     3781 tttttctgta tgtgagggcg gtctagctac agcattatta gctagaagtt atacagtcat
     3841 gactggttag gtgaaagcta ctttttcctt taccacttta taaatgggtt tttcctaaaa
     3901 ttatatgcac tgttgaggag gatgactatg taagatagcg cattctctgg ctaggaggaa
     3961 tgctgatgtt tgtataccct gctacaatat ggtttctctt tctctaaagt gatatatagc
     4021 cagatatttt tgtaataagt tgggagttga gtcctattaa atttagagaa agttatgatt
     4081 ccctcaaaga tgtaatacat actagatata gccaaaagag cgataaggac ctactgattg
     4141 tgatgttaat acatgataaa cattgaaaca ttttacattt taacagaaga gatgcctcag
     4201 attgaaacaa gagtgatatt ggttcaagaa gctggaaaac aagaagaact tataaaagcc
     4261 ttaaaggtac ggagttttag gttttagttt tttttgtaat gcaatttttc ttccatcagt
     4321 gtgtaaatag tttcatagct ttcttaacct catacttaag tctcttttcc acagactatt
     4381 aaaataatgg aagtccctgt tataaagata aaagaaagtt gtcctatctt tctagaagat
     4441 ctcctacaat attctcagct gccatggaaa atcgatgttc ttcttttatt ctctcaagat
     4501 tttcaggctg tatattaaaa cttatattaa gaactatgct aaccacctca tcaggaaccg
     4561 ttgtaggtgg cgtgggtttt cttggcaatc gactctcatg aaaactacga gctaaatatt
     4621 caatatgttc ctcttgacca actttattct gcattttttt tgaacgaggt ttagagcaag
     4681 cttcaggaaa ctgagacagg aattttatta aaaatttaaa ttttgaagaa agttcagggt
     4741 taatagcatc cattttttgc tttgcaagtt cctcagcatt cttaacaaaa gacgtctctt
     4801 ttgacatgtt taaagtttaa acctcctgtg tgaaattatt atccgctcat aattccacac
     4861 attatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     4921 acattaattg cgttgcgctc actgccaatt gctttccagt cgggaaacct gtcgtgccag
     4981 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     5041 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     5101 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     5161 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     5221 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     5281 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     5341 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     5401 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     5461 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     5521 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     5581 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     5641 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     5701 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     5761 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaa
//
