LOCUS       sequence-429494-        7754 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of human CALR wt.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7754)
  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 7754)
  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 wt
FEATURES             Location/Qualifiers
     source          1..7754
                     /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    2715..3264
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2859..2876)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3086..3105
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(3267..3269,3270..3272,3273..3986)
                     /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(3267..3269,3270..3272,3273..3983)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3312..3333)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3573..3592)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3920..3941
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             4148..4662
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    4824..4840
                     /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(4848..4854,4855..4872,4873..4878))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4893..4914
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5031..5048)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5202..5790)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5282..5301)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5961..6752,6753..6821))
                     /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     6584..6603
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6822..6926)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6994..7012
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7050..7072)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7172..7191
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7385..7407
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7535..7554
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             join(7754,1..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 aggaggatga
     2581 tgaggacaaa gatgaggatg aggaggatga ggaggacaag gaggaagatg aggaggaaga
     2641 tgtcccgggc caggccaagg acgagctgta ggaattcgtt aacctcgagc gggatcaatt
     2701 ccgccccccc cctaacgtta ctggccgaag ccgcttggaa taaggccggt gtgcgtttgt
     2761 ctatatgtta ttttccacca tattgccgtc ttttggcaat gtgagggccc ggaaacctgg
     2821 ccctgtcttc ttgacgagca ttcctagggg tctttcccct ctcgccaaag gaatgcaagg
     2881 tctgttgaat gtcgtgaagg aagcagttcc tctggaagct tcttgaagac aaacaacgtc
     2941 tgtagcgacc ctttgcaggc agcggaaccc cccacctggc gacaggtgcc tctgcggcca
     3001 aaagccacgt gtataagata cacctgcaaa ggcggcacaa ccccagtgcc acgttgtgag
     3061 ttggatagtt gtggaaagag tcaaatggct ctcctcaagc gtattcaaca aggggctgaa
     3121 ggatgcccag aaggtacccc attgtatggg atctgatctg gggcctcggt gcacatgctt
     3181 tacatgtgtt tagtcgaggt taaaaaaacg tctaggcccc ccgaaccacg gggacgtggt
     3241 tttcctttga aaaacacgat aataccatgg tgagcaaggg cgaggagctg ttcaccgggg
     3301 tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg
     3361 gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg
     3421 gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctacggc gtgcagtgct
     3481 tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag
     3541 gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg
     3601 aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca
     3661 aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc cacaacgtct
     3721 atatcatggc cgacaagcag aagaacggca tcaaggtgaa cttcaagatc cgccacaaca
     3781 tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg
     3841 gccccgtgct gctgcccgac aaccactacc tgagcaccca gtccgccctg agcaaagacc
     3901 ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggatcactc
     3961 tcggcatgga cgagctgtac aagtaaagcg gccgcgactc tagagtcgac ctgcaggcat
     4021 gcaagcttca ggtagccggc taacgttaac aaccggtacc tctagaacta tagctagcat
     4081 gcgcaaattt aaagcgctga tatcgataaa ataaaagatt ttatttagtc tccagaaaaa
     4141 ggggggaatg aaagacccca cctgtaggtt tggcaagcta gcttaagtaa cgccattttg
     4201 caaggcatgg aaaatacata actgagaata gagaagttca gatcaaggtt aggaacagag
     4261 agacagcaga atatgggcca aacaggatat ctgtggtaag cagttcctgc cccggctcag
     4321 ggccaagaac agatggtccc cagatgcggt cccgccctca gcagtttcta gagaaccatc
     4381 agatgtttcc agggtgcccc aaggacctga aatgaccctg tgccttattt gaactaacca
     4441 atcagttcgc ttctcgcttc tgttcgcgcg cttctgctcc ccgagctcaa taaaagagcc
     4501 cacaacccct cactcggcgc gccagtcctc cgatagactg cgtcgcccgg gtacccgtgt
     4561 atccaataaa ccctcttgca gttgcatccg acttgtggtc tcgctgttcc ttgggagggt
     4621 ctcctctgag tgattgacta cccgtcagcg ggggtctttc atgggtaaca gtttcttgaa
     4681 gttggagaac aacattctga gggtaggagt cgaatattaa gtaatcctga ctcaattagc
     4741 cactgttttg aatccacata ctccaatact cctgaaatag ttcattatgg acagcgcaga
     4801 aagagctggg gagaattgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     4861 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     4921 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     4981 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     5041 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     5101 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     5161 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     5221 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     5281 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     5341 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     5401 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     5461 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     5521 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     5581 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     5641 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     5701 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     5761 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     5821 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     5881 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     5941 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     6001 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     6061 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     6121 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     6181 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     6241 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     6301 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     6361 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     6421 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     6481 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     6541 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     6601 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     6661 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     6721 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     6781 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     6841 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     6901 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa
     6961 gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag gccctttcgt
     7021 ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc
     7081 acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt
     7141 gttggcgggt gtcggggctg gcttaactat gcggcatcag agcagattgt actgagagtg
     7201 caccatatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg catcaggcgc
     7261 cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc ctcttcgcta
     7321 ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat taagttgggt aacgccaggg
     7381 ttttcccagt cacgacgttg taaaacgacg gcgcaaggaa tggtgcatgc aaggagatgg
     7441 cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca
     7501 tgagcccgaa gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag
     7561 caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggcgattag
     7621 tccaatttgt taaagacagg atatcagtgg tccaggctct agttttgact caacaatatc
     7681 accagctgaa gcctatagag tacgagccat agataaaata aaagatttta tttagtctcc
     7741 agaaaaaggg ggga
//
