LOCUS       sequence-401319-        6076 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tardigrade (Hypsibius exemplaris) beta-tubulin Tub2D2 tagged with 
            mScarlet for expression in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6076)
  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 6076)
  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-Tub2D2-mScarlet
FEATURES             Location/Qualifiers
     source          1..6076
                     /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      2037..2046
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2043..2738
                     /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(2176..2194)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2444..2463
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    2864..2985
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2901..2920)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2955..2974
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2992..3447)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3129..3150)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3341..3360
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3474..3578
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3580..3937
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3580..3771
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3603..3623)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3788..3923
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3972..4766
                     /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             3972..4766
                     /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(4026..4045)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4636..4655
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4954..4973)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4998..5045
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5374..5962
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5863..5882
                     /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 gctagcgcca
      601 ccatgctcgg actctctacg ccttcagtac tgcgtcttcc cgttccttcg accatgcgag
      661 aaatcgtcca cctccaggcc ggccaatgcg gcaaccaaat cggctccaag ttctgggaaa
      721 ccatcaacgg cgagcatgga atcgacccca cgggatctta cacgggcgac aacgccaacc
      781 agcttgagag aatcgatgtt tactaccaga atgccagcag tgggaaatat gtgccaaggg
      841 ccgtgttggt ggaccttgaa ccgggcacga tggactcgat ccggtcgtcc gagtacggcc
      901 agctcttccg tccggataac tttgttttcg gccaaagtgg ggcgggcaac aactgggcta
      961 agggccatta cacggaggga gccgacctca tcgacacggt gatcgatgtc gtccgcaagg
     1021 aggccgagca ctgcgactgc ctgcagggct tccagctgac ccactcgctc ggcggtggaa
     1081 ccggatccgg catgggcacc ctcctcatct ccaagatccg cgaggagtat cccgatcgca
     1141 tcatgaccac attcagtgtc gttccctcgc ccaaggtgtc cgacaccgtg gtggagccgt
     1201 acaacgccac cctgtcagtc caccagctgg ttgagaacac cgacgagaca ttctgcatcg
     1261 acaacgaggc cctctacgac atctgcttcc gcaccctcaa gctgtcaacc ccggcgtacg
     1321 gcgacctgaa ccatctcgtc tcggcgacca tgtccggcgt gacgacctgc ttccgcttcc
     1381 ccggccagtt gaacgccgat ctccgcaaac tggccgtcaa catggtcccc ttcccgcggc
     1441 tgcacttctt catgccaggc tttgcgccgc tgactgctcg cggctctcaa acctaccgcg
     1501 ccatgtccgt gtcggagttg acccagcaga tgttcgactc gaagaacatg atgaccgcct
     1561 gcgatccccg ccacggacgc tacctcaccg tggctgccat cttccgcggc cggatgagca
     1621 tgaaggaggt cgatgataat atgttcaacg tgcagaacaa aaatagcagc catttcgttg
     1681 agtggatccc caacaacgtc aagaccgccg tctgcgatat ccccccgcgc ggactgtcca
     1741 tgtcgggcac cttcatcggc aacagcacgg ccattcagga gatcttcaag cgtatcggcg
     1801 agcagttcac cgccatgttc cgccgcaagg ccttcctcca ttggtacact ggcgaaggca
     1861 tggacgagat ggaattcacc gaagccgagt ccaacatgat ggacctggtg tcggaatacc
     1921 agcagtacca ggaggcgact gttgatgacg gcgcggaatt cgacgaggat acggaaaatg
     1981 tcgtggtcaa tggacatgaa aacgagaccg ccggaggtgg tggaggttca ccggtcgcca
     2041 ccatggtgag caagggcgag gcagtgatca aggagttcat gcggttcaag gtgcacatgg
     2101 agggctccat gaacggccac gagttcgaga tcgagggcga gggcgagggc cgcccctacg
     2161 agggcaccca gaccgccaag ctgaaggtga ccaagggtgg ccccctgccc ttctcctggg
     2221 acatcctgtc ccctcagttc atgtacggct ccagggcctt caccaagcac cccgccgaca
     2281 tccccgacta ctataagcag tccttccccg agggcttcaa gtgggagcgc gtgatgaact
     2341 tcgaggacgg cggcgccgtg accgtgaccc aggacacctc cctggaggac ggcaccctga
     2401 tctacaaggt gaagctccgc ggcaccaact tccctcctga cggccccgta atgcagaaga
     2461 agacaatggg ctgggaagcg tccaccgagc ggttgtaccc cgaggacggc gtgctgaagg
     2521 gcgacattaa gatggccctg cgcctgaagg acggcggccg ctacctggcg gacttcaaga
     2581 ccacctacaa ggccaagaag cccgtgcaga tgcccggcgc ctacaacgtc gaccgcaagt
     2641 tggacatcac ctcccacaac gaggactaca ccgtggtgga acagtacgaa cgctccgagg
     2701 gccgccactc caccggcggc atggacgagc tgtacaagta aagcggccgc gactctagat
     2761 cataatcagc cataccacat ttgtagaggt tttacttgct ttaaaaaacc tcccacacct
     2821 ccccctgaac ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc
     2881 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     2941 actgcattct agttgtggtt tgtccaaact catcaatgta tcttaaggcg taaattgtaa
     3001 gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
     3061 aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga
     3121 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
     3181 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt
     3241 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
     3301 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
     3361 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
     3421 cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt tcggggaaat gtgcgcggaa
     3481 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     3541 cctgataaat gcttcaataa tattgaaaaa ggaagagtcc tgaggcggaa agaaccagct
     3601 gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     3661 gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc
     3721 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac
     3781 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     3841 aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta
     3901 gtgaggaggc ttttttggag gcctaggctt ttgcaaagat cgatcaagag acaggatgag
     3961 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     4021 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     4081 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     4141 tgaatgaact gcaagacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
     4201 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     4261 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     4321 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     4381 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     4441 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcga
     4501 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     4561 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     4621 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
     4681 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     4741 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc
     4801 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     4861 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     4921 ggagttcttc gcccacccta gggggaggct aactgaaaca cggaaggaga caataccgga
     4981 aggaacccgc gctatgacgg caataaaaag acagaataaa acgcacggtg ttgggtcgtt
     5041 tgttcataaa cgcggggttc ggtcccaggg ctggcactct gtcgataccc caccgagacc
     5101 ccattggggc caatacgccc gcgtttcttc cttttcccca ccccaccccc caagttcggg
     5161 tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc cctgccatag cctcaggtta
     5221 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     5281 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     5341 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     5401 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     5461 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     5521 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     5581 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     5641 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     5701 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     5761 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     5821 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     5881 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     5941 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     6001 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     6061 tattaccgcc atgcat
//
