title: [Application of High-throughput Sequencing in Acute Myeloid Leukemia Patients with Positive FLT3-ITD].
abstract: OBJECTICE: To evaluate the application of high-throughput sequencing to sequence the FMS-like Tyrosine Kinase 3 internal tandem duplication (FLT3-ITD) in de novo acute myeloid leukemia (AML) patients with lower allelic ratio FLT3-ITD mutation or more than one ITD, and to analyze the feature of ITD.
  
  
  METHODS
  The genomic DNA of 23 AML patients with positive FLT3-ITD was amplified by PCR, capillary electrophoresis was used to detect the ITD mutation. Then, the FLT3 gene was amplified using primer with different barcode, and the product was analyzed by illumina Miseq, and the results were compared with UCSC database.
  
  
  RESULTS
  Out of 23 AML patients, 17 had a single ITD, and 3 had 2 ITDs, and the remaining 3 had 3 ITD detected by capillary electrophoresis. The high-throughput sequencing showed that 17 ITD were the complate duplications of wild-type FLT3, and the remaining 16 ITD were partial duplications in the all 33 ITDs. The same length ITD mutation contained 2 different ITD sequences in one patient with more than one ITD, and the other patient with 2 ITD had the same ITD insertion position. The ITD occurred in the regions from p. Y572 to p. L602 of the FLT3 protein, and all the patient ITD covered one or more amino acid between p. V592 and p. E598.
  
  
  CONCLUSIONS
  Illumina Miseq can analyze the sequence of ITDs precisely and accurately. ITD mutation varies widely, but the hotspots are concentrated.
authors.affiliation: Department of Clinical Laboratorial Examination, China-Japan Friendship Hospital, Beijing 100029, China.
authors.forename: Liang
authors.initials: L
authors.lastname: Ma
authors.affiliation: Department of Clinical Laboratorial Examination, China-Japan Friendship Hospital, Beijing 100029, China.
authors.forename: Yong-Wei
authors.initials: YW
authors.lastname: Jiang
authors.affiliation: Department of Clinical Laboratorial Examination, China-Japan Friendship Hospital, Beijing 100029, China.
authors.forename: Shao-Ting
authors.initials: ST
authors.lastname: Wang
authors.affiliation: Department of Clinical Laboratorial Examination, China-Japan Friendship Hospital, Beijing 100029, China.
authors.forename: Qian
authors.initials: Q
authors.lastname: Liu
authors.affiliation: Department of Clinical Laboratorial Examination, China-Japan Friendship Hospital, Beijing 100029, China.
authors.forename: Xiao
authors.initials: X
authors.lastname: Cong
authors.affiliation: Department of Clinical Laboratorial Examination, China-Japan Friendship Hospital, Beijing 100029, China.
authors.forename: Jun
authors.initials: J
authors.lastname: Shen
authors.affiliation: Department of Clinical Laboratorial Examination, China-Japan Friendship Hospital, Beijing 100029, China.
authors.forename: Yong-Yue
authors.initials: YY
authors.lastname: Cao
authors.affiliation: Department of Clinical Laboratorial Examination, China-Japan Friendship Hospital, Beijing 100029, China. E-mail: caoyongtong92@sina.com.
authors.forename: Yong-Tong
authors.initials: YT
authors.lastname: Cao
mesh_terms: D000483:Alleles; D004252:DNA Mutational Analysis; D005838:Genotype; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D015470:Leukemia, Myeloid, Acute / Q000235:genetics*; D009154:Mutation; D016133:Polymerase Chain Reaction; D020080:Tandem Repeat Sequences; D051941:fms-Like Tyrosine Kinase 3 / Q000235:genetics*
chemical_list: D051941:fms-Like Tyrosine Kinase 3