LOCUS       sequence-407207-        6218 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of human CALR ins5 (del18-197)-FLAG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6218)
  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 6218)
  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 (del18-197)-FLAG
FEATURES             Location/Qualifiers
     source          1..6218
                     /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             1714..1737
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(1809..1826)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1815..2039
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2085..2513
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2172..2191)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2382..2403
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2522..2542)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2527..2856
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2707..2842
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2769..2788
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2923..3717
                     /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(2977..2996)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3587..3606
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3891..4012
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3928..3947)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3982..4001
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4061..4077)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4061..4077)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4074..4096)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4085..4101
                     /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(4109..4115,4116..4133,4134..4139))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4154..4175
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4292..4309)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4463..5051)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4543..4562)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5222..6013,6014..6082))
                     /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     5845..5864
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6083..6187)
                     /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 cgccgacgat
     1021 tgggacttcc tgccacccaa gaagataaag gatcctgatg cttcaaaacc ggaagactgg
     1081 gatgagcggg ccaagatcga tgatcccaca gactccaagc ctgaggactg ggacaagccc
     1141 gagcatatcc ctgaccctga tgctaagaag cccgaggact gggatgaaga gatggacgga
     1201 gagtgggaac ccccagtgat tcagaaccct gagtacaagg gtgagtggaa gccccggcag
     1261 atcgacaacc cagattacaa gggcacttgg atccacccag aaattgacaa ccccgagtat
     1321 tctcccgatc ccagtatcta tgcctatgat aactttggcg tgctgggcct ggacctctgg
     1381 caggtcaagt ctggcaccat ctttgacaac ttcctcatca ccaacgatga ggcatacgct
     1441 gaggagtttg gcaacgagac gtggggcgta acaaaggcag cagagaaaca aatgaaggac
     1501 aaacaggacg aggagcagag gcttaaggag gaggaagaag acaagaaacg caaagaggag
     1561 gaggaggcag aggacaattg tcggaggatg atgaggacaa agatgaggat gaggaggatg
     1621 aggaggacaa ggaggaagat gaggaggaag atgtccccgg ccaggccaag gacgagctgt
     1681 agagaggcct gcctccaggg ctggactgag gccgactaca aggacgatga tgacaagtga
     1741 attctgcaga tatccagcac agtggcggcc gctcgagtct agagggcccg tttaaacccg
     1801 ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     1861 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     1921 tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag
     1981 caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc
     2041 ttctgaggcg gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg
     2101 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     2161 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     2221 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     2281 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     2341 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     2401 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     2461 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attaattctg
     2521 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     2581 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     2641 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     2701 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     2761 atttttttta tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag
     2821 tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc
     2881 attttcggat ctgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga
     2941 ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa
     3001 cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt
     3061 ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg
     3121 ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa
     3181 gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac
     3241 cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt
     3301 gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact
     3361 cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg
     3421 ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg
     3481 acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc
     3541 atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt
     3601 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc
     3661 gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg
     3721 ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg
     3781 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     3841 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta
     3901 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     3961 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     4021 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     4081 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     4141 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     4201 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     4261 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     4321 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     4381 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     4441 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     4501 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     4561 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     4621 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     4681 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     4741 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     4801 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     4861 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     4921 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     4981 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     5041 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     5101 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     5161 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     5221 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     5281 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     5341 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     5401 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     5461 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     5521 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     5581 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     5641 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     5701 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     5761 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     5821 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     5881 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     5941 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     6001 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     6061 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     6121 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     6181 ttccgcgcac atttccccga aaagtgccac ctgacgtc
//
