LOCUS       sequence-401320-        6002 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tardigrade (Hypsibius exemplaris) beta-tubulin Tub2D3 tagged with 
            mScarlet for expression in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6002)
  AUTHORS   Novotna Floriancicova K, Baltzis A, Smejkal J, Czernekova M, 
            Kaczmarek L, Maly J, Notredame C, Vinopal S
  TITLE     Phylogenetic and functional characterization of water bears 
            (Tardigrada) tubulins.
  JOURNAL   Sci Rep. 2023 Mar 30;13(1):5194. doi: 10.1038/s41598-023-31992-z.
  PUBMED    36997657
REFERENCE   2  (bases 1 to 6002)
  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: "Sci Rep."; 
            date: "2023-03-30"; volume: "13(1):5194. doi"; pages: " 
            10.1038/s41598-023-31992-z"
COMMENT     Sequence Label: ptHe-Tub2D3-mScarlet
FEATURES             Location/Qualifiers
     source          1..6002
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1963..1972
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1969..2664
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(2102..2120)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2370..2389
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    2790..2911
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2827..2846)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2881..2900
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2918..3373)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3055..3076)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3267..3286
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3400..3504
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3506..3863
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3506..3697
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3529..3549)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3714..3849
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3898..4692
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3898..4692
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3952..3971)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4562..4581
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4880..4899)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4924..4971
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5300..5888
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5789..5808
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc ggccaccatg
      601 cgtgaaatcg tccatttgca agcgggccaa tgtggaaacc aaatcggagc gaagttttgg
      661 gaagtcatct cggacgagca tggcatcgat cccaccggca cgtaccacgg agactccgat
      721 ctccagttgg agaggatcaa cgtttattac aatgaagcca ccggagggaa atatgtgccg
      781 cgcgcggttc tggtggatct cgagccggga acgatggact ctgtgcggtc cgggccattt
      841 ggacagcttt tccgtccgga caactttatt ttcggcctga gcggagccgg aaacaactgg
      901 gccaagggtc actacaccga gggcgccgaa ctggtggact cggtcctcga cgtggtccgc
      961 aaggaggccg agagctgcga ctgtctgcag ggcttccagc tgactcactc tctcggcggt
     1021 ggaaccggat cgggaatggg caccctgatg atctccaaga tccgtgagga gtatcccgat
     1081 cgcatcatga tgaccttcag tgtcgtccca tctcccaagg tgtccgacac tgtggtggag
     1141 ccgtacaacg ccaccctgtc cgtccaccag ctggtggaga acacggacga gaccttctgc
     1201 atcgacaatg aggcgctcta cgacatctgc ttccgcactc tcaagctgac cacaccaacg
     1261 tacggcgact tgaaccatct cgtctcggcg accatgtccg gtgtgactac ctgcctgcgc
     1321 ttcccgggtc agctgaacgc cgatctccgc aaactggccg tcaacatggt ccccttcccg
     1381 cggctgcact tcttcatgcc cggcttcgcc ccgctgacct cgcgtggttc ccagcagtac
     1441 cgcgcgctga ccgtgccgga actgacccag cagatgttcg acgcgaagaa tatgatggcc
     1501 gcctgcgatc cccgccacgg acgctacctc accgtggcgg caatcttccg cggccggatg
     1561 agcatgaagg aggtggatga acagctactg aacatccaga acaagaacag cagctacttc
     1621 gtcgagtgga tccccaacaa cgtgaagacc gcggtctgcg acattccacc gcgcggtttg
     1681 aagatgtccg ccaccttcat cggaaacagc actgccatcc aggagctgtt caaacgcgtt
     1741 tcggaacagt tcactgccat gttccgccga aaagcgttcc tgcattggta cactggagag
     1801 ggtatggacg agatggaatt caccgaagcc gagtccaaca tgaatgactt ggtgtcggag
     1861 taccaacagt accaggaggc aaccgctgaa gacgagggcg aattcgagga ggaaggcgag
     1921 ggtggagaag ctgaggctgg aggtggtgga ggttcaccgg tcgccaccat ggtgagcaag
     1981 ggcgaggcag tgatcaagga gttcatgcgg ttcaaggtgc acatggaggg ctccatgaac
     2041 ggccacgagt tcgagatcga gggcgagggc gagggccgcc cctacgaggg cacccagacc
     2101 gccaagctga aggtgaccaa gggtggcccc ctgcccttct cctgggacat cctgtcccct
     2161 cagttcatgt acggctccag ggccttcacc aagcaccccg ccgacatccc cgactactat
     2221 aagcagtcct tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc
     2281 gccgtgaccg tgacccagga cacctccctg gaggacggca ccctgatcta caaggtgaag
     2341 ctccgcggca ccaacttccc tcctgacggc cccgtaatgc agaagaagac aatgggctgg
     2401 gaagcgtcca ccgagcggtt gtaccccgag gacggcgtgc tgaagggcga cattaagatg
     2461 gccctgcgcc tgaaggacgg cggccgctac ctggcggact tcaagaccac ctacaaggcc
     2521 aagaagcccg tgcagatgcc cggcgcctac aacgtcgacc gcaagttgga catcacctcc
     2581 cacaacgagg actacaccgt ggtggaacag tacgaacgct ccgagggccg ccactccacc
     2641 ggcggcatgg acgagctgta caagtaaagc ggccgcgact ctagatcata atcagccata
     2701 ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga
     2761 aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca
     2821 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     2881 gtggtttgtc caaactcatc aatgtatctt aaggcgtaaa ttgtaagcgt taatattttg
     2941 ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc
     3001 ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt
     3061 tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc
     3121 tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg
     3181 tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga
     3241 aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg
     3301 ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg
     3361 ctacagggcg cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     3421 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     3481 caataatatt gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg aatgtgtgtc
     3541 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     3601 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     3661 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     3721 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     3781 atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt
     3841 ttggaggcct aggcttttgc aaagatcgat caagagacag gatgaggatc gtttcgcatg
     3901 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     3961 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     4021 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa
     4081 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     4141 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     4201 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     4261 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     4321 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     4381 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgagcat gcccgacggc
     4441 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     4501 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     4561 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     4621 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     4681 gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc
     4741 catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     4801 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     4861 accctagggg gaggctaact gaaacacgga aggagacaat accggaagga acccgcgcta
     4921 tgacggcaat aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt cataaacgcg
     4981 gggttcggtc ccagggctgg cactctgtcg ataccccacc gagaccccat tggggccaat
     5041 acgcccgcgt ttcttccttt tccccacccc accccccaag ttcgggtgaa ggcccagggc
     5101 tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc aggttactca tatatacttt
     5161 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     5221 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     5281 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     5341 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     5401 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     5461 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     5521 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     5581 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     5641 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     5701 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     5761 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     5821 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     5881 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     5941 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgccatgc
     6001 at
//
