LOCUS       sequence-258974-        5263 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pClimDCLifeAct:mCh actin filaments marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5263)
  AUTHORS   Kozyczkowska A, Najle SR, Ocana-Pallares E, Areste C, Shabardina V, 
            Ara PS, Ruiz-Trillo I, Casacuberta E
  TITLE     Stable transfection in protist Corallochytrium limacisporum 
            identifies novel cellular features among unicellular animals 
            relatives.
  JOURNAL   Curr Biol. 2021 Sep 27;31(18):4104-4110.e5. doi: 
            10.1016/j.cub.2021.06.061. Epub 2021 Jul 21.
  PUBMED    34293333
REFERENCE   2  (bases 1 to 5263)
  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: 
            "10.1016/j.cub.2021.06.061"; journalName: "Curr Biol"; date: 
            "2021-09-27- 27"; volume: "31"; issue: "18"; pages: "4104-4110.e5"
COMMENT     Sequence Label: Clim-DC2-lifeact-mCherry
FEATURES             Location/Qualifiers
     source          1..5263
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             569..619
                     /codon_start=1
                     /product="peptide that binds filamentous actin (Riedl et 
                     al., 2008)"
                     /label=Lifeact
                     /note="first 17 amino acids of budding yeast Abp140"
                     /translation="MGVADLIKKFESISKEE"
     CDS             635..1345
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(780..798)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1048..1067
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1242..1265
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     polyA_signal    1370..1491
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     1528..1547
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     CDS             join(1676..1744,1745..2536)
                     /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     complement(1894..1913)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2707..3295
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3196..3215
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3449..3466
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3583..3604
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3619..3624,3625..3642,3643..3649)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3657..3673
                     /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)."
     primer_bind     3662..3684
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3681..3697
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3681..3697
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             4486..5085
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4486..4505)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4982..5002
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5108..5229
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
ORIGIN
        1 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc gaaatgctgg
       61 gcgccttgaa actgaacctg ttcggatccg ggttgcgcgc ttccattgtt ttggtttccg
      121 tgtctgcaag gcgggtctgg acgtgacgct ttgggggaga gggagagcag agatacaatt
      181 ctagccacga atttcagcca atagtcacag aggggagttg cgatcctgcg aattggctta
      241 ggtctgtgag gtctttcacg gtttcgaaag cagcgcagag gggagaccga ctaccaaaag
      301 ccataacact ccaccgagat atccgcactc ttcgctagat gagcttgctt ctcctcttgt
      361 cacaaatcag cccaatgata gtcactgttc aacaacggtg cagtgtaaat attcgttgtg
      421 cgtccttgat gagcagaagg tacgagaagc aggaaatcat ttcataatgg aaaaccattc
      481 actgtctttc tgtattgcga aaagagtgat cataggtaat cgcaacggcg ggtctttaga
      541 acagtcacat accagatcaa gcttggggat gggtgtcgca gatttgatca agaaattcga
      601 aagcatctca aaggaagaag ggactagtga tatcatggtg agcaagggcg aggaggacaa
      661 catggccatc atcaaggagt tcatgcgctt caaggtgcac atggagggct ccgtgaacgg
      721 ccacgagttc gagatcgagg gcgagggcga gggccgcccc tacgagggca cccagaccgc
      781 caagctgaag gtgaccaagg gcggccccct gcccttcgcc tgggacatcc tgtcccctca
      841 gttcatgtac ggctccaagg cctacgtgaa gcaccccgcc gacatccccg actacttgaa
      901 gctgtccttc cccgagggct tcaagtggga gcgcgtgatg aacttcgagg acggcggcgt
      961 ggtgaccgtg acccaggact cctccctgca ggacggcgag ttcatctaca aggtgaagct
     1021 gcgcggcacc aacttcccct ccgacggccc cgtaatgcag aagaagacca tgggctggga
     1081 ggcctcctcc gagcggatgt accccgagga cggcgccctg aagggcgaga tcaagcagag
     1141 gctgaagctg aaggacggcg gccactacga cgccgaggtc aagaccacct acaaggccaa
     1201 gaagcccgtg cagctgcccg gcgcctacaa cgtcaacatc aagctggaca tcacctccca
     1261 caacgaggac tacaccatcg tggaacagta cgagcgcgcc gagggccgcc actccaccgg
     1321 cggcatggac gagctgtaca agtgacatat gctagcagat ctagggccct aagatacatt
     1381 gatgagtttg gacaaaccac aacaagaatg cagtgaaaaa aatgctttat ttgtgaaatt
     1441 tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt tgccggctgg
     1501 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     1561 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     1621 gcatcgcctt ctatcgcctt cttgacgagt tcttctgaat tgaaaaagga agagtatgag
     1681 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     1741 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     1801 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     1861 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     1921 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     1981 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     2041 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     2101 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     2161 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     2221 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     2281 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     2341 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     2401 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     2461 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     2521 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     2581 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     2641 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     2701 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     2761 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     2821 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     2881 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     2941 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     3001 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     3061 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     3121 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     3181 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     3241 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     3301 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     3361 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     3421 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg
     3481 cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga
     3541 caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga gttagctcac
     3601 tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt
     3661 gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca agcttatcgg
     3721 tgggactgtg agaactgcta cctgcctgtg cagaggagag cacactacat tcaccattga
     3781 aaatatggcc cactgaactg atcaagccaa gtgaagactg cgcgcctgtg ttctgattct
     3841 tcccatggta caagtactga aactttggaa gcgccagaga attcacagac gcacagaaga
     3901 cagcgcatgg tggaggctgt gtgtgttcac cgaggtgtcc tctgcggaag gtgccgttca
     3961 ttatgtattc ggagtaaagc acaacctctt caatgctgct cccttccttc gcttgtttct
     4021 tggatagtta tcgtctgtca gaatctttcg gtacattcag tatagagttc gcgatcggta
     4081 gcaacagtcg gacggaccct gtcaggacgg tgtattcggt ggcctacttt tcttttgttg
     4141 catagctgtt cctgacgcga tgcgccaagc gccgtcatat tgtgctgtgt gcccctatgt
     4201 ggagcaggca tagagctctt gccaaatgtt gggaagagct gttggagaag cgacagagca
     4261 gtgccaagac gacgaacgag gtgtgcgagc tgtagtgcta gtgcttccta ggttctggtt
     4321 ttactgacac ctatttgaag gaataatttc tgctcacaca cagttttgta accgctcact
     4381 attgtgttac agctcatcat aagctactat aacacatctt tgcggcttta agctagtgta
     4441 ttcatctaat ttacaatcat gcgcgagtgt atctctattc ccgggatgac cgagtacaag
     4501 cccacggtgc gcctcgccac ccgcgacgac gtccccaggg ccgtacgcac cctcgccgcc
     4561 gcgttcgccg actaccccgc cacgcgccac accgtcgatc cggaccgcca catcgagcgg
     4621 gtcaccgagc tgcaagaact cttcctcacg cgcgtcgggc tcgacatcgg caaggtgtgg
     4681 gtcgcggacg acggcgccgc ggtggcggtc tggaccacgc cggagagcgt cgaagcgggg
     4741 gcggtgttcg ccgagatcgg cccgcgcatg gccgagttga gcggttcccg gctggccgcg
     4801 cagcaacaga tggaaggcct cctggcgccg caccggccca aggagcccgc gtggttcctg
     4861 gccaccgtcg gcgtctcgcc cgaccaccag ggcaagggtc tgggcagcgc cgtcgtgctc
     4921 cccggagtgg aggcggccga gcgcgccggg gtgcccgcct tcctggagac ctccgcgccc
     4981 cgcaacctcc ccttctacga gcggctcggc ttcaccgtca ccgccgacgt cgaggtgccc
     5041 gaaggaccgc gcacctggtg catgacccgc aagcccggtg cctgacctcg agcatcgata
     5101 gacatgataa gatacattga tgagtttgga caaaccacaa caagaatgca gtgaaaaaaa
     5161 tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat
     5221 aaacaagtta acaacaacaa ttctagaggg cctgcttgcc gaa
//
