LOCUS       sequence-429498-        7793 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of human CALR ins5.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7793)
  AUTHORS   Masubuchi N, Araki M, Yang Y, Hayashi E, Imai M, Edahiro Y, Hironaka
            Y, Mizukami Y, Kihara Y, Takei H, Nudejima M, Koike M, Ohsaka A, 
            Komatsu N
  TITLE     Mutant calreticulin interacts with MPL in the secretion pathway for 
            activation on the cell surface.
  JOURNAL   Leukemia. 2020 Feb;34(2):499-509. doi: 10.1038/s41375-019-0564-z. 
            Epub 2019 Aug 28.
  PUBMED    31462733
REFERENCE   2  (bases 1 to 7793)
  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: 
            "10.1038/s41375-019-0564-z"; journalName: "Leukemia"; date: 
            "2020-02"; volume: "34"; issue: "2"; pages: "499-509"
COMMENT     Sequence Label: pMSCV-IRES-GFP/CALR ins5
FEATURES             Location/Qualifiers
     source          1..7793
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    580..921
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             988..1404
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1334..1356
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1372..1388
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    2754..3303
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2898..2915)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3125..3144
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(3306..3308,3309..3311,3312..4025)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(3306..3308,3309..3311,3312..4022)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3351..3372)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3612..3631)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3959..3980
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             4187..4701
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    4863..4879
                     /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)."
     promoter        complement(join(4887..4893,4894..4911,4912..4917))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4932..4953
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5070..5087)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5241..5829)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5321..5340)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6000..6791,6792..6860))
                     /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     6623..6642
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6861..6965)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7033..7051
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7089..7111)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7211..7230
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7424..7446
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7574..7593
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             join(7793,1..516)
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
ORIGIN
        1 atgaaagacc ccacctgtag gtttggcaag ctagcttaag taacgccatt ttgcaaggca
       61 tggaaaatac ataactgaga atagagaagt tcagatcaag gttaggaaca gagagacagc
      121 agaatatggg ccaaacagga tatctgtggt aagcagttcc tgccccggct cagggccaag
      181 aacagatggt ccccagatgc ggtcccgccc tcagcagttt ctagagaacc atcagatgtt
      241 tccagggtgc cccaaggacc tgaaatgacc ctgtgcctta tttgaactaa ccaatcagtt
      301 cgcttctcgc ttctgttcgc gcgcttctgc tccccgagct caataaaaga gcccacaacc
      361 cctcactcgg cgcgccagtc ctccgataga ctgcgtcgcc cgggtacccg tattcccaat
      421 aaagcctctt gctgtttgca tccgaatcgt ggactcgctg atccttggga gggtctcctc
      481 agattgattg actgcccacc tcgggggtct ttcatttgga ggttccaccg agatttggag
      541 acccctgccc agggaccacc gacccccccg ccgggaggta agctggccag cggtcgtttc
      601 gtgtctgtct ctgtctttgt gcgtgtttgt gccggcatct aatgtttgcg cctgcgtctg
      661 tactagttag ctaactagct ctgtatctgg cggacccgtg gtggaactga cgagttctga
      721 acacccggcc gcaaccctgg gagacgtccc agggactttg ggggccgttt ttgtggcccg
      781 acctgaggaa gggagtcgat gtggaatccg accccgtcag gatatgtggt tctggtagga
      841 gacgagaacc taaaacagtt cccgcctccg tctgaatttt tgctttcggt ttggaaccga
      901 agccgcgcgt cttgtctgct gcagcgctgc agcatcgttc tgtgttgtct ctgtctgact
      961 gtgtttctgt atttgtctga aaattagggc cagactgtta ccactccctt aagtttgacc
     1021 ttaggtcact ggaaagatgt cgagcggatc gctcacaacc agtcggtaga tgtcaagaag
     1081 agacgttggg ttaccttctg ctctgcagaa tggccaacct ttaacgtcgg atggccgcga
     1141 gacggcacct ttaaccgaga cctcatcacc caggttaaga tcaaggtctt ttcacctggc
     1201 ccgcatggac acccagacca ggtcccctac atcgtgacct gggaagcctt ggcttttgac
     1261 ccccctccct gggtcaagcc ctttgtacac cctaagcctc cgcctcctct tcctccatcc
     1321 gccccgtctc tcccccttga acctcctcgt tcgaccccgc ctcgatcctc cctttatcca
     1381 gccctcactc cttctctagg cgccggaatt cgccaccatg ctgctatccg tgccgctgct
     1441 gctcggcctc ctcggcctgg ccgtcgccga gcctgccgtc tacttcaagg agcagtttct
     1501 ggacggagac gggtggactt cccgctggat cgaatccaaa cacaagtcag attttggcaa
     1561 attcgttctc agttccggca agttctacgg tgacgaggag aaagataaag gtttgcagac
     1621 aagccaggat gcacgctttt atgctctgtc ggccagtttc gagcctttca gcaacaaagg
     1681 ccagacgctg gtggtgcagt tcacggtgaa acatgagcag aacatcgact gtgggggcgg
     1741 ctatgtgaag ctgtttccta atagtttgga ccagacagac atgcacggag actcagaata
     1801 caacatcatg tttggtcccg acatctgtgg ccctggcacc aagaaggttc atgtcatctt
     1861 caactacaag ggcaagaacg tgctgatcaa caaggacatc cgttgcaagg atgatgagtt
     1921 tacacacctg tacacactga ttgtgcggcc agacaacacc tatgaggtga agattgacaa
     1981 cagccaggtg gagtccggct ccttggaaga cgattgggac ttcctgccac ccaagaagat
     2041 aaaggatcct gatgcttcaa aaccggaaga ctgggatgag cgggccaaga tcgatgatcc
     2101 cacagactcc aagcctgagg actgggacaa gcccgagcat atccctgacc ctgatgctaa
     2161 gaagcccgag gactgggatg aagagatgga cggagagtgg gaacccccag tgattcagaa
     2221 ccctgagtac aagggtgagt ggaagccccg gcagatcgac aacccagatt acaagggcac
     2281 ttggatccac ccagaaattg acaaccccga gtattctccc gatcccagta tctatgccta
     2341 tgataacttt ggcgtgctgg gcctggacct ctggcaggtc aagtctggca ccatctttga
     2401 caacttcctc atcaccaacg atgaggcata cgctgaggag tttggcaacg agacgtgggg
     2461 cgtaacaaag gcagcagaga aacaaatgaa ggacaaacag gacgaggagc agaggcttaa
     2521 ggaggaggaa gaagacaaga aacgcaaaga ggaggaggag gcagaggaca attgtcggag
     2581 gatgatgagg acaaagatga ggatgaggag gatgaggagg acaaggagga agatgaggag
     2641 gaagatgtcc ccggccaggc caaggacgag ctgtagagag gcctgcctcc agggctggac
     2701 tgaggcctga gaattcgtta acctcgagcg ggatcaattc cgcccccccc ctaacgttac
     2761 tggccgaagc cgcttggaat aaggccggtg tgcgtttgtc tatatgttat tttccaccat
     2821 attgccgtct tttggcaatg tgagggcccg gaaacctggc cctgtcttct tgacgagcat
     2881 tcctaggggt ctttcccctc tcgccaaagg aatgcaaggt ctgttgaatg tcgtgaagga
     2941 agcagttcct ctggaagctt cttgaagaca aacaacgtct gtagcgaccc tttgcaggca
     3001 gcggaacccc ccacctggcg acaggtgcct ctgcggccaa aagccacgtg tataagatac
     3061 acctgcaaag gcggcacaac cccagtgcca cgttgtgagt tggatagttg tggaaagagt
     3121 caaatggctc tcctcaagcg tattcaacaa ggggctgaag gatgcccaga aggtacccca
     3181 ttgtatggga tctgatctgg ggcctcggtg cacatgcttt acatgtgttt agtcgaggtt
     3241 aaaaaaacgt ctaggccccc cgaaccacgg ggacgtggtt ttcctttgaa aaacacgata
     3301 ataccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     3361 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     3421 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     3481 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     3541 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     3601 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     3661 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     3721 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     3781 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     3841 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     3901 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     3961 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     4021 agtaaagcgg ccgcgactct agagtcgacc tgcaggcatg caagcttcag gtagccggct
     4081 aacgttaaca accggtacct ctagaactat agctagcatg cgcaaattta aagcgctgat
     4141 atcgataaaa taaaagattt tatttagtct ccagaaaaag gggggaatga aagaccccac
     4201 ctgtaggttt ggcaagctag cttaagtaac gccattttgc aaggcatgga aaatacataa
     4261 ctgagaatag agaagttcag atcaaggtta ggaacagaga gacagcagaa tatgggccaa
     4321 acaggatatc tgtggtaagc agttcctgcc ccggctcagg gccaagaaca gatggtcccc
     4381 agatgcggtc ccgccctcag cagtttctag agaaccatca gatgtttcca gggtgcccca
     4441 aggacctgaa atgaccctgt gccttatttg aactaaccaa tcagttcgct tctcgcttct
     4501 gttcgcgcgc ttctgctccc cgagctcaat aaaagagccc acaacccctc actcggcgcg
     4561 ccagtcctcc gatagactgc gtcgcccggg tacccgtgta tccaataaac cctcttgcag
     4621 ttgcatccga cttgtggtct cgctgttcct tgggagggtc tcctctgagt gattgactac
     4681 ccgtcagcgg gggtctttca tgggtaacag tttcttgaag ttggagaaca acattctgag
     4741 ggtaggagtc gaatattaag taatcctgac tcaattagcc actgttttga atccacatac
     4801 tccaatactc ctgaaatagt tcattatgga cagcgcagaa agagctgggg agaattgtga
     4861 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     4921 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     4981 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     5041 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     5101 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     5161 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     5221 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     5281 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     5341 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     5401 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     5461 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     5521 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     5581 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     5641 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
     5701 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     5761 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     5821 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     5881 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     5941 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     6001 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     6061 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     6121 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     6181 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     6241 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     6301 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     6361 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     6421 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     6481 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     6541 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     6601 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     6661 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     6721 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     6781 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     6841 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     6901 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     6961 ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca
     7021 ttaacctata aaaataggcg tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac
     7081 ggtgaaaacc tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat
     7141 gccgggagca gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg
     7201 cttaactatg cggcatcaga gcagattgta ctgagagtgc accatatgcg gtgtgaaata
     7261 ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc attcgccatt caggctgcgc
     7321 aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg
     7381 ggatgtgctg caaggcgatt aagttgggta acgccagggt tttcccagtc acgacgttgt
     7441 aaaacgacgg cgcaaggaat ggtgcatgca aggagatggc gcccaacagt cccccggcca
     7501 cggggcctgc caccataccc acgccgaaac aagcgctcat gagcccgaag tggcgagccc
     7561 gatcttcccc atcggtgatg tcggcgatat aggcgccagc aaccgcacct gtggcgccgg
     7621 tgatgccggc cacgatgcgt ccggcgtaga ggcgattagt ccaatttgtt aaagacagga
     7681 tatcagtggt ccaggctcta gttttgactc aacaatatca ccagctgaag cctatagagt
     7741 acgagccata gataaaataa aagattttat ttagtctcca gaaaaagggg gga
//
