LOCUS       sequence-405247-        5661 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  The vector contains 1kbp of the upstream and downstream regions of 
            the cahs3 gene from Ramazzottius varieornatus, with the 
            RvCAHS3-mEGFP gene positioned in between..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5661)
  AUTHORS   Tanaka S, Aoki K, Arakawa K
  TITLE     In vivo expression vector derived from anhydrobiotic tardigrade 
            genome enables live imaging in Eutardigrada.
  JOURNAL   Proc Natl Acad Sci U S A. 2023 Jan 31;120(5):e2216739120. doi: 
            10.1073/pnas.2216739120. Epub 2023 Jan 24.
  PUBMED    36693101
REFERENCE   2  (bases 1 to 5661)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2023-01-31"; volume: "120(5):e2216739120. 
            doi"; pages: " 10.1073/pnas.2216739120. Epub 2023 Jan 24"
COMMENT     Sequence Label: pRvCAHS3-CAHS3-mEGFP
FEATURES             Location/Qualifiers
     source          1..5661
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             911..925
                     /codon_start=1
                     /product="enterokinase recognition and cleavage site"
                     /label=enterokinase site
                     /translation="DDDDK"
     CDS             join(1954..1956,1957..1959,1960..2673)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELSR"
     primer_bind     complement(1999..2020)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2260..2279)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2607..2628
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3746..3763)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3917..4505)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3997..4016)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4676..5467,5468..5536))
                     /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     5299..5318
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5537..5641)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacctgggtc gatcgacatt gattattgac tagttgttct cgggaatcca ccccctcgat
       61 ttctacgaat tcttctttgt aaaatcccca aggaccatct tcttgctttc ttttccggta
      121 ctgatagctc aaaggtgctg ctgcatagtg gccgtgttgc cggtgcgatg gcggagctga
      181 ccgtgtgata aaaaaatcgt aaaataatcc tggttaaaca tgcatcacta ttattctaaa
      241 actcttcctt cttcccgctt tttgtgtttc aagtggatct tgagcaaaaa aaacaaaaga
      301 ctttgttact ggtgctggac atcagccaag cttcaaaagt cgcgttactt acatacgaaa
      361 ttcacaattt tgacgcaaaa aacggcaaat ttccactgaa agacctatat aataacgtcc
      421 acttcgttga attgtttgct gacgttctta gtgttcgaag agaatatttc ggtggaatgc
      481 ggttttcttt ggcaatatgc gcgaaaaacc tttattctgt ttgctattcg cacctatcga
      541 tagtgaacga cacatctctt gtaaacatcg ccttcaacct atacgggctg ttattcttct
      601 tcttaagtaa tgagtcaagc tttggaaata ccagacggct cgccgctcga gtcgacaagc
      661 atattctcat gacatattgc cgtctgtacg acaggccact cgcattcttt tccaggcgta
      721 cagccggttt ttgccgtgga ttaactctca taggacctct cacgctggtg atatcaggaa
      781 atcacaccac acaagctgtt gttatcgcgc tttgatgatc actacctgta caactcgcag
      841 cattttccgt cgggaaaaag cctggcagaa agttcagctg agcacacaag acgacgacga
      901 cgacgacgac gacgacgacg ataaaacttg ccgactgtgt cagttcagct tcacactatt
      961 cccgtgtact ttgcagcact tacccagctc ttgtttattc tttcgcagcc tttcgctaac
     1021 gccctgtcgc aaaccatgtc ttcccgacag aaccagcaat cgtccagcca acactcgtcc
     1081 tccagccagc aaggtggtca aggtggtcaa ggtgttcaag gaagttccag ctactcgcgc
     1141 accgaggtcc acaccagcag tggaggacct accatcggtg gagcccagcg aactgtcccc
     1201 gtcccccctg gatctcactc cgaggtccat gaggagcgtg aggtcatcaa gcatggtacc
     1261 aaaaccgaaa gcgagaccca cgtcgtcacc gtcccagtga caactttcgg cagcaccaac
     1321 atggaatctg tccgaaccgg cttcaccgtc acccaagaca agaacttgac cgttgctgct
     1381 cccaacatcg ctgctcccat ccacagcaac ctcgacctta acctcggcgg tggagctcgc
     1441 gctgaaatta ccgcagggac caccgttgac ttgagcaaga tccagcgcaa ggatttggga
     1501 cctgaagagt atgctcgcta caaggccaag gtcgagcaac tggccaggca agatgagcaa
     1561 gacgcgggta tgcgcgctgc ccagtaccga gaggaagtgg agcgtgatgc cgaactcatc
     1621 cgacagatct tggagcgaca acacatccgt gatcttgaat tccgcaagga aatggttgag
     1681 aaccaagtca accgacaaga gagagaaatc cagctggagg ctgagtacgc aatgcgagcc
     1741 ctcgagcttg agcgcaatgc cgctaaagag gctttggaaa gcgccaaggc ccagactaac
     1801 gtcaatgtca aggtcgagtc cgctattggt accaccgtct ccaagggtgc aatccagact
     1861 tccgccgaca agagcagcac caccaagact ggacccacca ccgtcactca gattaaacat
     1921 accgaacaac acactgaacg ccgaggcggc cgcatggtga gcaagggcga ggagctgttc
     1981 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc
     2041 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc
     2101 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg
     2161 cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg
     2221 cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc
     2281 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc
     2341 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac
     2401 aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc
     2461 cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc
     2521 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc caagcttagc
     2581 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg
     2641 atcactctcg gcatggacga gctgtctaga taaatttgct cataaccaca ccatcataca
     2701 atttttcacc atttctaaaa agaaaatgtt ttcgtccagt cttttgtgca ctctgaaatg
     2761 tcatatcatg aataacttca gttttagcgc cgcaaggaaa atgtgagacg gttaatatca
     2821 cgaacgccgg taaacaatat tcgtggtcaa taaaacgatt aatttctaca tcagtcatgc
     2881 tttcttccgt gcactgcttt aaacaaggca aaccgatttc taacggaaga atgtactatt
     2941 acctctgctg aactgtggcc cccacgactt ttctttgtgg aatttaccgt aaacgttgag
     3001 tgcggtactc aatattcaag aaaatactcc atgcgtgcac atattacgta agaatgactt
     3061 ctcgaccatt gtctaaaagt cactggcgca tatgtgcagt accctgacca ttgcaagaca
     3121 ggcagattgc atgctgtgct ggtgatcgcc cagggctgtt ttttgtttac tgtcacagac
     3181 cgcgactgcc ttcgcaatga ctgatccctc ccaccgaagt aaacataatt atgaccacaa
     3241 aatcgagttt tattacatgt cacaaaagtg gctgcagaag atctcacaga ttatgacaac
     3301 tcatcgtaaa attattatca aagttcaaat tcgcggagca aaatggctca ccagataact
     3361 atgctaagat cagacttggt agcatactca gtggatatgt cttgtcaaag tggggtccac
     3421 ggattccgca tttgtcacag tagttgtgcg tgtggtggtc acaccagtgg tcagatcgct
     3481 gcctacgccc cctgtagtcg tcccttgagt catcccaagg actctttcat ccggaaccaa
     3541 atcggcttga cgcttgtctt tgaaattcat gccggggtcc tcgccacctt ggtcttcgcg
     3601 gacatctaca gaaacgacgt ttccgtctcc gtcgacatgc gccttctcca cccgctcata
     3661 ctttacctcc tttggatccg ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     3721 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     3781 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     3841 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     3901 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     3961 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     4021 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     4081 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     4141 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     4201 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     4261 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     4321 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     4381 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     4441 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     4501 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     4561 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     4621 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     4681 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     4741 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     4801 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     4861 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     4921 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     4981 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     5041 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     5101 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     5161 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     5221 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     5281 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     5341 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     5401 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     5461 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     5521 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     5581 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     5641 gcacatttcc ccgaaaagtg c
//
