LOCUS       sequence-417225-        9032 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Retroviral expression of human CALR del52-FLAG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9032)
  AUTHORS   Araki M, Yang Y, Masubuchi N, Hironaka Y, Takei H, Morishita S, 
            Mizukami Y, Kan S, Shirane S, Edahiro Y, Sunami Y, Ohsaka A, Komatsu
            N
  TITLE     Activation of the thrombopoietin receptor by mutant calreticulin in 
            CALR-mutant myeloproliferative neoplasms.
  JOURNAL   Blood. 2016 Jan 27. pii: blood-2015-09-671172.
  PUBMED    26817954
REFERENCE   2  (bases 1 to 9032)
  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: "Blood. 2016
            Jan 27. pii: blood-2015-09-671172."
COMMENT     Sequence Label: pMSCV-IRES-GFP/CALR del52-FLAG
FEATURES             Location/Qualifiers
     source          1..9032
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..517
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
     misc_feature    581..922
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             989..1405
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1335..1357
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1373..1389
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     CDS             2652..2675
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             3924..3947
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     misc_feature    3994..4543
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4138..4155)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4365..4384
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(4546..4548,4549..4551,4552..5265)
                     /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(4546..4548,4549..4551,4552..5262)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4591..4612)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4852..4871)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5199..5220
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             5427..5941
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    6103..6119
                     /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(6127..6133,6134..6151,6152..6157))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6172..6193
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6310..6327)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6481..7069)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6561..6580)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7240..8031,8032..8100))
                     /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     7863..7882
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8101..8205)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     8273..8291
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(8329..8351)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     8451..8470
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8664..8686
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     8814..8833
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 aatgaaagac cccacctgta ggtttggcaa gctagcttaa gtaacgccat tttgcaaggc
       61 atggaaaata cataactgag aatagagaag ttcagatcaa ggttaggaac agagagacag
      121 cagaatatgg gccaaacagg atatctgtgg taagcagttc ctgccccggc tcagggccaa
      181 gaacagatgg tccccagatg cggtcccgcc ctcagcagtt tctagagaac catcagatgt
      241 ttccagggtg ccccaaggac ctgaaatgac cctgtgcctt atttgaacta accaatcagt
      301 tcgcttctcg cttctgttcg cgcgcttctg ctccccgagc tcaataaaag agcccacaac
      361 ccctcactcg gcgcgccagt cctccgatag actgcgtcgc ccgggtaccc gtattcccaa
      421 taaagcctct tgctgtttgc atccgaatcg tggactcgct gatccttggg agggtctcct
      481 cagattgatt gactgcccac ctcgggggtc tttcatttgg aggttccacc gagatttgga
      541 gacccctgcc cagggaccac cgaccccccc gccgggaggt aagctggcca gcggtcgttt
      601 cgtgtctgtc tctgtctttg tgcgtgtttg tgccggcatc taatgtttgc gcctgcgtct
      661 gtactagtta gctaactagc tctgtatctg gcggacccgt ggtggaactg acgagttctg
      721 aacacccggc cgcaaccctg ggagacgtcc cagggacttt gggggccgtt tttgtggccc
      781 gacctgagga agggagtcga tgtggaatcc gaccccgtca ggatatgtgg ttctggtagg
      841 agacgagaac ctaaaacagt tcccgcctcc gtctgaattt ttgctttcgg tttggaaccg
      901 aagccgcgcg tcttgtctgc tgcagcgctg cagcatcgtt ctgtgttgtc tctgtctgac
      961 tgtgtttctg tatttgtctg aaaattaggg ccagactgtt accactccct taagtttgac
     1021 cttaggtcac tggaaagatg tcgagcggat cgctcacaac cagtcggtag atgtcaagaa
     1081 gagacgttgg gttaccttct gctctgcaga atggccaacc tttaacgtcg gatggccgcg
     1141 agacggcacc tttaaccgag acctcatcac ccaggttaag atcaaggtct tttcacctgg
     1201 cccgcatgga cacccagacc aggtccccta catcgtgacc tgggaagcct tggcttttga
     1261 cccccctccc tgggtcaagc cctttgtaca ccctaagcct ccgcctcctc ttcctccatc
     1321 cgccccgtct ctcccccttg aacctcctcg ttcgaccccg cctcgatcct ccctttatcc
     1381 agccctcact ccttctctag gcgccggaat tcgccaccat gctgctatcc gtgccgctgc
     1441 tgctcggcct cctcggcctg gccgtcgccg agcctgccgt ctacttcaag gagcagtttc
     1501 tggacggaga cgggtggact tcccgctgga tcgaatccaa acacaagtca gattttggca
     1561 aattcgttct cagttccggc aagttctacg gtgacgagga gaaagataaa ggtttgcaga
     1621 caagccagga tgcacgcttt tatgctctgt cggccagttt cgagcctttc agcaacaaag
     1681 gccagacgct ggtggtgcag ttcacggtga aacatgagca gaacatcgac tgtgggggcg
     1741 gctatgtgaa gctgtttcct aatagtttgg accagacaga catgcacgga gactcagaat
     1801 acaacatcat gtttggtccc gacatctgtg gccctggcac caagaaggtt catgtcatct
     1861 tcaactacaa gggcaagaac gtgctgatca acaaggacat ccgttgcaag gatgatgagt
     1921 ttacacacct gtacacactg attgtgcggc cagacaacac ctatgaggtg aagattgaca
     1981 acagccaggt ggagtccggc tccttggaag acgattggga cttcctgcca cccaagaaga
     2041 taaaggatcc tgatgcttca aaaccggaag actgggatga gcgggccaag atcgatgatc
     2101 ccacagactc caagcctgag gactgggaca agcccgagca tatccctgac cctgatgcta
     2161 agaagcccga ggactgggat gaagagatgg acggagagtg ggaaccccca gtgattcaga
     2221 accctgagta caagggtgag tggaagcccc ggcagatcga caacccagat tacaagggca
     2281 cttggatcca cccagaaatt gacaaccccg agtattctcc cgatcccagt atctatgcct
     2341 atgataactt tggcgtgctg ggcctggacc tctggcaggt caagtctggc accatctttg
     2401 acaacttcct catcaccaac gatgaggcat acgctgagga gtttggcaac gagacgtggg
     2461 gcgtaacaaa ggcagcagag aaacaaatga aggacaaaca ggacgaggag cagaggacaa
     2521 ggaggatgat gaggacaaag atgaggatga ggaggatgag gaggacaagg aggaagatga
     2581 ggaggaagat gtccccggcc aggccaagga cgagctgtag agaggcctgc ctccagggct
     2641 ggactgaggc cgactacaaa gacgatgacg ataaatagga attcgccacc atgctgctat
     2701 ccgtgccgct gctgctcggc ctcctcggcc tggccgtcgc cgagcctgcc gtctacttca
     2761 aggagcagtt tctggacgga gacgggtgga cttcccgctg gatcgaatcc aaacacaagt
     2821 cagattttgg caaattcgtt ctcagttccg gcaagttcta cggtgacgag gagaaagata
     2881 aaggtttgca gacaagccag gatgcacgct tttatgctct gtcggccagt ttcgagcctt
     2941 tcagcaacaa aggccagacg ctggtggtgc agttcacggt gaaacatgag cagaacatcg
     3001 actgtggggg cggctatgtg aagctgtttc ctaatagttt ggaccagaca gacatgcacg
     3061 gagactcaga atacaacatc atgtttggtc ccgacatctg tggccctggc accaagaagg
     3121 ttcatgtcat cttcaactac aagggcaaga acgtgctgat caacaaggac atccgttgca
     3181 aggatgatga gtttacacac ctgtacacac tgattgtgcg gccagacaac acctatgagg
     3241 tgaagattga caacagccag gtggagtccg gctccttgga agacgattgg gacttcctgc
     3301 cacccaagaa gataaaggat cctgatgctt caaaaccgga agactgggat gagcgggcca
     3361 agatcgatga tcccacagac tccaagcctg aggactggga caagcccgag catatccctg
     3421 accctgatgc taagaagccc gaggactggg atgaagagat ggacggagag tgggaacccc
     3481 cagtgattca gaaccctgag tacaagggtg agtggaagcc ccggcagatc gacaacccag
     3541 attacaaggg cacttggatc cacccagaaa ttgacaaccc cgagtattct cccgatccca
     3601 gtatctatgc ctatgataac tttggcgtgc tgggcctgga cctctggcag gtcaagtctg
     3661 gcaccatctt tgacaacttc ctcatcacca acgatgaggc atacgctgag gagtttggca
     3721 acgagacgtg gggcgtaaca aaggcagcag agaaacaaat gaaggacaaa caggacgagg
     3781 agcagaggac aaggaggatg atgaggacaa agatgaggat gaggaggatg aggaggacaa
     3841 ggaggaagat gaggaggaag atgtccccgg ccaggccaag gacgagctgt agagaggcct
     3901 gcctccaggg ctggactgag gccgactaca aagacgatga cgataaatag gaattcgtta
     3961 acctcgagcg ggatcaattc cgcccccccc ctaacgttac tggccgaagc cgcttggaat
     4021 aaggccggtg tgcgtttgtc tatatgttat tttccaccat attgccgtct tttggcaatg
     4081 tgagggcccg gaaacctggc cctgtcttct tgacgagcat tcctaggggt ctttcccctc
     4141 tcgccaaagg aatgcaaggt ctgttgaatg tcgtgaagga agcagttcct ctggaagctt
     4201 cttgaagaca aacaacgtct gtagcgaccc tttgcaggca gcggaacccc ccacctggcg
     4261 acaggtgcct ctgcggccaa aagccacgtg tataagatac acctgcaaag gcggcacaac
     4321 cccagtgcca cgttgtgagt tggatagttg tggaaagagt caaatggctc tcctcaagcg
     4381 tattcaacaa ggggctgaag gatgcccaga aggtacccca ttgtatggga tctgatctgg
     4441 ggcctcggtg cacatgcttt acatgtgttt agtcgaggtt aaaaaaacgt ctaggccccc
     4501 cgaaccacgg ggacgtggtt ttcctttgaa aaacacgata ataccatggt gagcaagggc
     4561 gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc
     4621 cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg
     4681 aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg
     4741 acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc
     4801 aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc
     4861 aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag
     4921 ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac
     4981 tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac
     5041 ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag
     5101 aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcacccag
     5161 tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg
     5221 accgccgccg ggatcactct cggcatggac gagctgtaca agtaaagcgg ccgcgactct
     5281 agagtcgacc tgcaggcatg caagcttcag gtagccggct aacgttaaca accggtacct
     5341 ctagaactat agctagcatg cgcaaattta aagcgctgat atcgataaaa taaaagattt
     5401 tatttagtct ccagaaaaag gggggaatga aagaccccac ctgtaggttt ggcaagctag
     5461 cttaagtaac gccattttgc aaggcatgga aaatacataa ctgagaatag agaagttcag
     5521 atcaaggtta ggaacagaga gacagcagaa tatgggccaa acaggatatc tgtggtaagc
     5581 agttcctgcc ccggctcagg gccaagaaca gatggtcccc agatgcggtc ccgccctcag
     5641 cagtttctag agaaccatca gatgtttcca gggtgcccca aggacctgaa atgaccctgt
     5701 gccttatttg aactaaccaa tcagttcgct tctcgcttct gttcgcgcgc ttctgctccc
     5761 cgagctcaat aaaagagccc acaacccctc actcggcgcg ccagtcctcc gatagactgc
     5821 gtcgcccggg tacccgtgta tccaataaac cctcttgcag ttgcatccga cttgtggtct
     5881 cgctgttcct tgggagggtc tcctctgagt gattgactac ccgtcagcgg gggtctttca
     5941 tgggtaacag tttcttgaag ttggagaaca acattctgag ggtaggagtc gaatattaag
     6001 taatcctgac tcaattagcc actgttttga atccacatac tccaatactc ctgaaatagt
     6061 tcattatgga cagcgcagaa agagctgggg agaattgtga aattgttatc cgctcacaat
     6121 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
     6181 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     6241 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     6301 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     6361 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     6421 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     6481 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     6541 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     6601 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     6661 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     6721 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     6781 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     6841 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     6901 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     6961 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     7021 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     7081 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     7141 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     7201 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     7261 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     7321 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     7381 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     7441 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     7501 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     7561 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     7621 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     7681 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     7741 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     7801 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     7861 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     7921 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     7981 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     8041 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     8101 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     8161 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     8221 agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg
     8281 tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat
     8341 gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg
     8401 tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga
     8461 gcagattgta ctgagagtgc accatatgcg gtgtgaaata ccgcacagat gcgtaaggag
     8521 aaaataccgc atcaggcgcc attcgccatt caggctgcgc aactgttggg aagggcgatc
     8581 ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt
     8641 aagttgggta acgccagggt tttcccagtc acgacgttgt aaaacgacgg cgcaaggaat
     8701 ggtgcatgca aggagatggc gcccaacagt cccccggcca cggggcctgc caccataccc
     8761 acgccgaaac aagcgctcat gagcccgaag tggcgagccc gatcttcccc atcggtgatg
     8821 tcggcgatat aggcgccagc aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt
     8881 ccggcgtaga ggcgattagt ccaatttgtt aaagacagga tatcagtggt ccaggctcta
     8941 gttttgactc aacaatatca ccagctgaag cctatagagt acgagccata gataaaataa
     9001 aagattttat ttagtctcca gaaaaagggg gg
//
