LOCUS       sequence-401322-        6041 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tardigrade (Hypsibius exemplaris) gamma-tubulin Tub3 tagged with 
            mScarlet for expression in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6041)
  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 6041)
  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-Tub3-mScarlet
FEATURES             Location/Qualifiers
     source          1..6041
                     /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      2002..2011
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2008..2703
                     /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(2141..2159)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2409..2428
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    2829..2950
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2866..2885)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2920..2939
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2957..3412)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3094..3115)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3306..3325
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3439..3543
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3545..3902
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3545..3736
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3568..3588)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3753..3888
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3937..4731
                     /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             3937..4731
                     /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(3991..4010)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4601..4620
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4919..4938)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4963..5010
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5339..5927
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5828..5847
                     /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 cctcgctcca tcatctccct gcagctcggc cagtgcggca accaggtcgg catggaattc
      661 tggaagaaat tgtgcatgga gcatggcatc gagagcaacg gccagctgaa gaattccggc
      721 gcggagggcg actttcggga taggaaggat ttcttcttct accaggccga cgacagccac
      781 tacattccgc gcgccatcct catcgatctc gagccgcgtg tgattcagac catcaagaat
      841 tccgactacg gcaagctgtt taatccggaa aacatttatg tgtcggagca gggcggtggt
      901 gcgggaaaca actgggctgc aggattcgga atggggcaag acgtctcgga gaaagtgttc
      961 gacatgattg agcgtgaagc ggaaggctgc gacggattag agggcttcat tctcagtcac
     1021 tccattgccg gcggtaccgg gtccggcatg ggctcatacg tcctggagca tttgcatgat
     1081 cgtttcccga agaagctcat tcagaccttc tccatcttcc ccaatcagga cgactctgcc
     1141 agcgatgtgg tggttcagcc gtacaactcg atcttgacga tgcgccggct ggcggagaac
     1201 gccgactgtg tggtggtgct ggacaacacg gcgctcaatt cgatcgctgc agaacggctg
     1261 cgaatcgacc aaccgagctt gacagagatc aacaatatgg tctcaacggt gatggccgcc
     1321 agtacgtcca cattgcgcta tcccggttac atgtacaaca actttgtcgg cttgatggcc
     1381 ccgctcattc ccatgccgaa gctccatttt ctcatggccg gctatgctcc actgacgtcg
     1441 ttggaccaga cgagcggtgg catccagaaa acgacggtat ttgacgttat gcgtcgttta
     1501 cttcagccga agaacatgat gctgtccccg gggaaggatg ccctggctgc cgacaatcgc
     1561 tatctctcca ttttgaacat cattcagggc gaagtggaat ctggcgagat tcacaaaagc
     1621 ctgatgcgca ttaaagaacg caatctggtc aagtttatgg aatggggtcc ggccagcatc
     1681 catgtggccc agtcgaagcg ctcgccgttt gtcgcgtcca accaccgcat cagcggactg
     1741 atgttggcca accacaccag cgtgcaggga ctcttcgaca gcatcgtcaa gcagtacgac
     1801 aagatgaaga agcgcaatgc ctacttggat cgcttcacca atgagaagaa ttgggccggt
     1861 gtgtcggaca tggagcagtc gagggaggtg gtggccaacc tgattgagga gtacaaggca
     1921 gccgcgaaac cggattacat agactatctg cgggggaaac tgcagagtaa gaagatggga
     1981 ggtggtggag gttcaccggt cgccaccatg gtgagcaagg gcgaggcagt gatcaaggag
     2041 ttcatgcggt tcaaggtgca catggagggc tccatgaacg gccacgagtt cgagatcgag
     2101 ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa ggtgaccaag
     2161 ggtggccccc tgcccttctc ctgggacatc ctgtcccctc agttcatgta cggctccagg
     2221 gccttcacca agcaccccgc cgacatcccc gactactata agcagtcctt ccccgagggc
     2281 ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg ccgtgaccgt gacccaggac
     2341 acctccctgg aggacggcac cctgatctac aaggtgaagc tccgcggcac caacttccct
     2401 cctgacggcc ccgtaatgca gaagaagaca atgggctggg aagcgtccac cgagcggttg
     2461 taccccgagg acggcgtgct gaagggcgac attaagatgg ccctgcgcct gaaggacggc
     2521 ggccgctacc tggcggactt caagaccacc tacaaggcca agaagcccgt gcagatgccc
     2581 ggcgcctaca acgtcgaccg caagttggac atcacctccc acaacgagga ctacaccgtg
     2641 gtggaacagt acgaacgctc cgagggccgc cactccaccg gcggcatgga cgagctgtac
     2701 aagtaaagcg gccgcgactc tagatcataa tcagccatac cacatttgta gaggttttac
     2761 ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg
     2821 ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     2881 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     2941 atgtatctta aggcgtaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt
     3001 tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca
     3061 aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta
     3121 aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta
     3181 cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg
     3241 aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga
     3301 aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg
     3361 ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc
     3421 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     3481 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     3541 agtcctgagg cggaaagaac cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc
     3601 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt
     3661 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     3721 cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg
     3781 cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct
     3841 cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca
     3901 aagatcgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac
     3961 gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca
     4021 atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt
     4081 gtcaagaccg acctgtccgg tgccctgaat gaactgcaag acgaggcagc gcggctatcg
     4141 tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga
     4201 agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct
     4261 cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg
     4321 gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg
     4381 gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc
     4441 gaactgttcg ccaggctcaa ggcgagcatg cccgacggcg aggatctcgt cgtgacccat
     4501 ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac
     4561 tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt
     4621 gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct
     4681 cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc
     4741 tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca
     4801 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga
     4861 tcctccagcg cggggatctc atgctggagt tcttcgccca ccctaggggg aggctaactg
     4921 aaacacggaa ggagacaata ccggaaggaa cccgcgctat gacggcaata aaaagacaga
     4981 ataaaacgca cggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc
     5041 actctgtcga taccccaccg agaccccatt ggggccaata cgcccgcgtt tcttcctttt
     5101 ccccacccca ccccccaagt tcgggtgaag gcccagggct cgcagccaac gtcggggcgg
     5161 caggccctgc catagcctca ggttactcat atatacttta gattgattta aaacttcatt
     5221 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     5281 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     5341 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     5401 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     5461 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     5521 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     5581 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     5641 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     5701 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     5761 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     5821 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     5881 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     5941 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     6001 tatcccctga ttctgtggat aaccgtatta ccgccatgca t
//
