LOCUS       sequence-407198-        6425 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of human CALR ins5 (del198-308)-FLAG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6425)
  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 6425)
  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 (del198-308)-FLAG
FEATURES             Location/Qualifiers
     source          1..6425
                     /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             1921..1944
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(2016..2033)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2022..2246
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2292..2720
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2379..2398)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2589..2610
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2729..2749)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2734..3063
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2914..3049
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2976..2995
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3130..3924
                     /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(3184..3203)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3794..3813
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4098..4219
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4135..4154)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4189..4208
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4268..4284)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4268..4284)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4281..4303)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4292..4308
                     /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(4316..4322,4323..4340,4341..4346))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4361..4382
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4499..4516)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4670..5258)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4750..4769)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5429..6220,6221..6289))
                     /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     6052..6071
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6290..6394)
                     /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 ggaagataac
     1561 tttggcgtgc tgggcctgga cctctggcag gtcaagtctg gcaccatctt tgacaacttc
     1621 ctcatcacca acgatgaggc atacgctgag gagtttggca acgagacgtg gggcgtaaca
     1681 aaggcagcag agaaacaaat gaaggacaaa caggacgagg agcagaggct taaggaggag
     1741 gaagaagaca agaaacgcaa agaggaggag gaggcagagg acaattgtcg gaggatgatg
     1801 aggacaaaga tgaggatgag gaggatgagg aggacaagga ggaagatgag gaggaagatg
     1861 tccccggcca ggccaaggac gagctgtaga gaggcctgcc tccagggctg gactgaggcc
     1921 gactacaagg acgatgatga caagtgaatt ctgcagatat ccagcacagt ggcggccgct
     1981 cgagtctaga gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca
     2041 gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac
     2101 tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat
     2161 tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca
     2221 tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag
     2281 ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     2341 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     2401 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     2461 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
     2521 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     2581 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
     2641 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
     2701 acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc
     2761 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
     2821 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     2881 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     2941 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     3001 tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     3061 aaaaagctcc cgggagcttg tatatccatt ttcggatctg atcaagagac aggatgagga
     3121 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     3181 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     3241 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     3301 aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     3361 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     3421 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     3481 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     3541 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     3601 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc
     3661 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     3721 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     3781 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     3841 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     3901 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     3961 cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     4021 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     4081 agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     4141 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     4201 aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt
     4261 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     4321 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     4381 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     4441 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     4501 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     4561 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     4621 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     4681 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     4741 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     4801 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     4861 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     4921 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     4981 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     5041 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     5101 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     5161 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     5221 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     5281 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     5341 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     5401 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     5461 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     5521 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     5581 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     5641 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     5701 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     5761 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     5821 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     5881 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     5941 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     6001 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     6061 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     6121 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     6181 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     6241 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     6301 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     6361 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     6421 acgtc
//
