LOCUS       sequence-407205-        6061 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of human CALR ins5 (del198-430)-FLAG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6061)
  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 6061)
  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: pcDNA3.1/CALR ins5 (del198-430)-FLAG
FEATURES             Location/Qualifiers
     source          1..6061
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1555..1578
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(1652..1669)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1658..1882
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1928..2356
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2015..2034)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2225..2246
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2365..2385)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2370..2699
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2550..2685
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2612..2631
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2766..3560
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2820..2839)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3430..3449
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3734..3855
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3771..3790)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3825..3844
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3904..3920)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3904..3920)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3917..3939)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3928..3944
                     /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(3952..3958,3959..3976,3977..3982))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3997..4018
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4135..4152)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4306..4894)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4386..4405)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5065..5856,5857..5925))
                     /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     5688..5707
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5926..6030)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttggta ccgagctcgg atccactagt ccagtgtggt ggaattcgcc
      961 accatgctgc tatccgtgcc gctgctgctc ggcctcctcg gcctggccgt cgccgagcct
     1021 gccgtctact tcaaggagca gtttctggac ggagacgggt ggacttcccg ctggatcgaa
     1081 tccaaacaca agtcagattt tggcaaattc gttctcagtt ccggcaagtt ctacggtgac
     1141 gaggagaaag ataaaggttt gcagacaagc caggatgcac gcttttatgc tctgtcggcc
     1201 agtttcgagc ctttcagcaa caaaggccag acgctggtgg tgcagttcac ggtgaaacat
     1261 gagcagaaca tcgactgtgg gggcggctat gtgaagctgt ttcctaatag tttggaccag
     1321 acagacatgc acggagactc agaatacaac atcatgtttg gtcccgacat ctgtggccct
     1381 ggcaccaaga aggttcatgt catcttcaac tacaagggca agaacgtgct gatcaacaag
     1441 gacatccgtt gcaaggatga tgagtttaca cacctgtaca cactgattgt gcggccagac
     1501 aacacctatg aggtgaagat tgacaacagc caggtggagt ccggctcctt ggaagactac
     1561 aaagacgatg acgataaata ggaattctgc agatatccag cacagtggcg gccgctcgag
     1621 tctagagggc ccgtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca
     1681 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
     1741 ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg
     1801 gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct
     1861 ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg
     1921 tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     1981 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     2041 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     2101 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     2161 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     2221 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     2281 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     2341 aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
     2401 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
     2461 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     2521 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     2581 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg
     2641 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
     2701 agctcccggg agcttgtata tccattttcg gatctgatca agagacagga tgaggatcgt
     2761 ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc
     2821 tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc
     2881 tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg
     2941 aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag
     3001 ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg
     3061 ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg
     3121 caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac
     3181 atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg
     3241 acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc
     3301 ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg
     3361 aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc
     3421 aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc
     3481 gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc
     3541 ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc
     3601 caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg
     3661 aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt
     3721 cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     3781 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     3841 catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc
     3901 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     3961 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     4021 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     4081 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     4141 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     4201 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     4261 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     4321 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     4381 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     4441 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     4501 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     4561 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     4621 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     4681 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     4741 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     4801 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     4861 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     4921 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     4981 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     5041 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     5101 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     5161 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     5221 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     5281 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     5341 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     5401 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     5461 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     5521 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     5581 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     5641 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     5701 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     5761 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     5821 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     5881 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     5941 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     6001 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
     6061 c
//
