Cloning and sequencing of KIR2DL1 framework gene cDNA and identification of a novel allele.
- Author:
Ge SUN
1
,
2
;
Chang WANG
;
Jianxin ZHEN
;
Guobin ZHANG
;
Yunping XU
;
Zhihui DENG
Author Information
- Publication Type:Journal Article
- MeSH: Alleles; Base Sequence; China; Cloning, Molecular; DNA, Complementary; chemistry; genetics; Haplotypes; Humans; Male; Mutation, Missense; Receptors, KIR2DL1; genetics; Sequence Analysis, DNA; methods
- From: Chinese Journal of Medical Genetics 2016;33(5):694-697
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo develop an assay for cDNA cloning and haplotype sequencing of KIR2DL1 framework gene and determine the genotype of an ethnic Han from southern China.
METHODSTotal RNA was isolated from peripheral blood sample, and complementary DNA (cDNA) transcript was synthesized by RT-PCR. The entire coding sequence of the KIR2DL1 framework gene was amplified with a pair of KIR2DL1-specific PCR primers. The PCR products with a length of approximately 1.2 kb were then subjected to cloning and haplotype sequencing.
RESULTSA specific target fragment of the KIR2DL1 framework gene was obtained. Following allele separation, a wild-type KIR2DL1*00302 allele and a novel variant allele, KIR2DL1*031, were identified. Sequence alignment with KIR2DL1 alleles from the IPD-KIR Database showed that the novel allele KIR2DL1*031 has differed from the closest allele KIR2DL1*00302 by a non-synonymous mutation at CDS nt 188A>G (codon 42 GAG>GGG) in exon 4, which has caused an amino acid change Glu42Gly. The sequence of the novel allele KIR2DL1*031 was submitted to GenBank under the accession number KP025960 and to the IPD-KIR Database under the submission number IWS40001982. A name KIR2DL1*031 has been officially assigned by the World Health Organization (WHO) Nomenclature Committee.
CONCLUSIONAn assay for cDNA cloning and haplotype sequencing of KIR2DL1 has been established, which has a broad applications in KIR studies at allelic level.
