MNS, Duffy, and Kell blood groups among the Uygur population of

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MNS, Duffy, and Kell blood groups among
the Uygur population of Xinjiang, China
G.Y. Lin, X.L. Du, J.J. Shan, Y.N. Zhang Y.Q. Zhang and Q.H. Wang
Blood Transfusion Department, People’s Liberation Army 474th Hospital,
Urumqi, Xinjiang Uyghur Autonomous Region, China
Corresponding author: G.Y. Lin / Q.H. Wang
E-mail: [email protected] / [email protected]
Genet. Mol. Res. 16 (1): gmr16019176
Received September 6, 2016
Accepted February 14, 2017
Published March 15, 2017
DOI http://dx.doi.org/10.4238/gmr16019176
Copyright © 2017 The Authors. This is an open-access article distributed under the terms of
the Creative Commons Attribution ShareAlike (CC BY-SA) 4.0 License.
ABSTRACT. Human blood groups are a significant resource for
patients, leading to a fierce international competition in the screening
of rare blood groups. Some rare blood group screening programs
have been implemented in western countries and Japan, but not
particularly in China. Recently, the genetic background of ABO and
Rh blood groups for different ethnic groups or regions in China has
been focused on increasingly. However, rare blood groups such as MN,
Duffy, Kidd, MNS, and Diego are largely unexplored. No systematic
reports exist concerning the polymorphisms and allele frequencies
of rare blood groups in China’s ethnic minorities such as Uygur and
Kazak populations of Xinjiang, unlike those on the Han population.
Therefore, this study aimed to investigate the allele frequencies of rare
blood groups, namely, MNS, Duffy, Kell, Dombrock, Diego, Kidd,
Scianna, Colton, and Lutheran in the Uygur population of Xinjiang
Single specific primer-polymerase chain reaction was performed
for genotyping and statistical analysis of 9 rare blood groups in 158
Uygur individuals. Allele frequencies were compared with distribution
among other ethnic groups. Observed and expected values of genotype
frequencies were compared using the chi-square test. Genotype
frequencies obeyed the Hardy-Weinberg equilibrium (P > 0.5) and
Genetics and Molecular Research 16 (1): gmr16019176
G.Y. Lin et al.
2
allele frequencies were stable. Of all subjects detected, 4 cases carried
the rare phenotype S-s- of MNS blood group (frequency of 0.0253), and
1 case carried the phenotype Jka-b- (frequency of 0.0063). Frequencies
of the four groups, MNS, Duffy, Dombrock, and Diego, in the Uygur
population differed from those in other ethnic groups. Gene distribution
of the Kell, Kidd, and Colton was similar to that in Tibetan and Han
populations, though there were some discrepancies. Gene distribution
of Scianna and Lutheran groups showed monomorphism similar to
that in Tibetan and Han populations. These findings could contribute
to the investigation of the origin, evolution, and hematology of Uygur
population of Xinjiang and assist in screening of rare blood groups
in ethnic minorities, meeting of clinical blood supply demands, and
building of the national rare blood group library.
Key words: Uygur population; Rare blood group; Genotyping;
Allele frequency; PCR-SSP; Xinjiang
INTRODUCTION
Human blood is as valuable as oils and rare earth elements and considered an
integral part of national strategic resources (Yang et al., 2014). The International Society
of Blood Transfusion has officially recognized 33 human erythrocyte blood group systems,
which include over 300 inheritable blood group antigens (Ji et al., 2012). The international
competition in the screening of rare blood groups is fierce (Chen et al., 2014). While the
rare blood group screening programs have been implemented in western countries and Japan,
much has yet to be done to accomplish this task in China.
Frequencies of rare blood group antigens are believed to vary significantly with race
and ethnicity. For example, the frequency of RhD-negative blood types is 20% in individuals
of European-descent, 10% in equatorial races, 0.4% in Mongolians, and only 0.3% in China’s
Han ethnic group. In the MN blood group system, blood type M is dominant in North China,
while blood type N is dominant in South China (Shulman, 1990). This blood group is not
only associated with hemolytic transfusion reactions and hemolytic disease of the newborn,
but also with the occurrence of hypertension (Delanghe et al., 1995). The positive rate of
Mia antigen is as high as 24.7% among the Dai people in Xishuangbanna, Yunnan Province,
while the positive rate of Mia antigen is only 5.4% in the Han population of Shanghai. The
overall positive rate of Dia antigen in the Chinese population is as high as 9%. Association
of the hemolytic transfusion reaction and hemolytic disease of the newborn with the anti-Dia
antibody has been reported previously (Liu, 2011).
The Jk(a-b-) phenotype of the Kidd blood group has been reportedly found in the
Macao Blood Center (Liu, 2011). The other four phenotypes are Jk (a+b-), Jk (a-b+), Jk
(a+b+), and Jk (a-b-). The Jk (a-b-) phenotype is very rare in any population. The frequency
of the JK (a-b-) phenotype for the Chinese Han ethnic group is less than one in ten thousand
(Ji et al., 2012; Chen et al., 2014), therefore, it is usually difficult to find matched blood for
transfusions quickly.
Individuals with Fy(a-b-) phenotype of the Duffy blood group have been reported
in blood centers in Qingdao and Inner Mongolia (Wang et al., 1993). In the Chinese Han
Genetics and Molecular Research 16 (1): gmr16019176
Rare blood type genes in the Uygur population
3
ethnic group, the dominant phenotype is Fy(a+b-), accounting for 91%, while the Fy(a+b+)
phenotype accounts for only 9%.
The Kell blood group antigen K1 is the most common alloantibody. Caucasian (British,
American, etc.) Women, About 40% of women carrying the anti-Ki antibody will give birth
to babies with severe hemolysis. In Europe and the United States blood transfusion treatment
work, K original identification of blood transfusion routine items (Roychoudhury et al., 1988).
Therefore, rare blood groups represent one of the major research topics.
China is a vast territory with many ethnic groups, and the distribution of blood groups
vary significantly. For example, 13 ethnic groups have lived in Xinjiang for generations. The
distribution of rare blood groups in Xinjiang displays unique features because of relative
geographical isolation, interethnic marriages, and historical migration. Under ideal situations,
the allele frequencies of a given population would remain stable across generations. However, in
reality, allele frequencies of a population are constantly changing on account of gene mutation,
gene recombination, natural selection, migration, and genetic drift, resulting in the continuous
evolution within species. The evolution of a population can be characterized usually by allele
frequency. This study focused on the allele frequencies of nine rare blood groups in the Uygur
population of Xinjiang, including MNS, Duffy, Kidd, Diego, and Dombrock to understand
the genetic make-up and blood relationship within the Uygur ethnic group. We hope the data
obtained in the study can contribute to the analysis of the origin, fusion, and migration of the
Uygur ethnic group, enriching the hematological and genetic studies of China’s ethnic groups
and the human race. The findings can be used to prepare for emergencies and build the rare
blood group library.
MATERIAL AND METHODS
Subjects
A total of 158 inpatient and outpatient cases at the 474th Hospital of People’s
Liberation Army were considered for the study. The subjects were unrelated by blood for
three generations. There were 92 males and 66 females, and their ages ranged from 1-84
years old (median 50.81). The experiment conformed to the PRC code of ethics for clinical
trials. The experiment was approved by the hospital Ethics Committee (approval No.:
SQS20140501). Blood samples were collected after obtaining informed consent from all
study participants.
Sample collection
A total of 162 blood samples were collected between May 2014 to January 2015,
seven monthly. A volume of 3.5 mL of blood was drawn from the median cubital vein from
each subject and preserved with ethylenediaminetetraacetic acid (EDTA) anticoagulant. Four
blood samples were excluded because of the presence of the anticoagulant heparin as this
would affect DNA extraction. Thus, 158 samples were finally included. None of the subjects
had hematological diseases, and most of them received blood type identification before blood
withdrawal. The sample size, statistical design, and the types of kits used were determined by
reference to similar studies of other ethnic minorities in China (Zhao and Li, 2009; Zhang et
al., 2014).
Genetics and Molecular Research 16 (1): gmr16019176
G.Y. Lin et al.
4
Blood sample processing
The whole blood samples containing the heparin were excluded. The concentration of
the extracted DNA was 30-100 ng/mL. If the concentration was too high, the sample was diluted
with Tris-EDTA buffer. DNA was diluted with Tris-EDTA buffer to a final concentration of 30120 ng/mL, with the optimal concentration of 30-50 ng/mL. The 260/280 purity ratio was 1.601.80. The DNA samples must not be dissolved in EDTA with concentration above 0.5 mM. To
ensure the integrity of the samples, they were transported below 4°C. The DNA samples were
dissolved in 1X Tris-EDTA buffer (pH 8.0-9.0), and extracted genomic DNA samples were
preserved at -20°C for less than 1 year.
Reagents and equipment
Reagents
The polymerase chain reaction-sequence specific primer (PCR-SSP) blood typing kits
for MNS, Duffy, Kell, Dombrock, Diego, Kidd, Scianna, Colton and Lutheran blood groups
were manufactured by Tianjin Super Biotechnology Development Co., Ltd Tianjin city, in
China. The whole blood genomic DNA extraction kit (PROTRANS, Hockenheim, Germany)
and agarose (Biowest regular agarose G-10, Manufacturerd to specifications Distributed by
GENE COMPANYLTD, ORIGIN:SPAIN) Taq polymerase (5 U/mL) was manufactured by
Promega (Madison, WI, USA). Other reagents used were ethidium bromide (10 mg/mL,
analytically pure), 1X TBE buffer (each component being analytically pure), and purified
water (deionized water).
Equipment
Biosafety cabinet (Haier,Dalian city, in China); high-speed microcentrifuge TG16-W
(Changsha Weierkang Xiangying Centrifuge, in China); HC-2515 high-speed centrifuge
(USTC ZONKIA, Heifei city, in China); DYY-BC electrophoresis apparatus, electrophoresis
tank, and WD-9413B gel imaging analysis system (Beijing Liuyi Instrument Factory, in
China); BIO-RAD PCR instrument (model 580BR 6679, USA); continuous micro pipette (5100 mL); vortex mixer; Midea microwave oven to heat the liquid to 100°C.
Methods
Genomic DNA extraction
DNA extraction was performed in 17 steps according to the instruction of the
PROTRANS DNA Extraction Kit. Genomic DNA concentration and purity were determined
with an ultraviolet-visible (UV-Vis) spectrophotometer. The genomic DNA samples were
preserved at -20°C and detected in batches.
PCR-SSP typing
The specific primers were designed for the 9 rare blood groups via base substitution
Genetics and Molecular Research 16 (1): gmr16019176
5
Rare blood type genes in the Uygur population
or deletion according to the sequences in the Genebank (The primer blood typing kits for
MNS, Duffy, Kell, Dombrock, Diego, Kidd, Scianna, Colton and Lutheran blood groups were
manufactured by Tianjin Super Biotechnology Development Co., Ltd Tianjin city, in China).
Table 1. The 3,-terminal base of the sense primer was designed to allow amplification only
if the base matches the template exactly. The conserved fragment of human growth hormone
(HGH) gene was used to design the internal control primers. The internal control was added
into each sample well and in each run.
First, the extracted genomic DNA was adjusted to the optimal concentration of 30-50
ng/mL. PCR procedures were as follows: 96°C for 2 min, 1 cycle; 96°C for 20 s, 68°C for 60 s,
5 cycles; 96°C for 20 s, 65°C for 45 s, 72°C for 30 s, 10 cycles; 96°C for 20 s, 62°C for 45 s,
72°C for 30 s, 15 cycles; 72°C for 3 min, 1 cycle. The amplification products were preserved
at 4°C. The reaction mix consisted of the followings: dNTP-Buffer (440 mL condensed dNTPBuffer + 560 mL PCR-grade sterilized water = 1000 mL) with mixing and centrifugation. The
mixture of buffer-enzyme-samples was prepared at a certain proportion. For each subject, the
mixture was 220 mL dNTP-buffer + 1.7 mL Taq polymerase (5 U/mL) + 25 mL DNA sample
= 246.7 mL, with mixing and centrifugation. Into each well (1-22 wells), 10 mL of the above
mixture was added. Paraffin oil (15-20 mL) was also added into each well. The tube was
properly capped and placed into the pre-configured PCR instrument. PCR programs were run
according to the above steps. In the meantime, 2.5% agarose gel was prepared.
Table 1. Base pairs, sites of nucleotide mutation, and primers for the typing of the 9 rare blood groups.
Rare blood group
MNS
Duffy
Kell
Dombrock
Diego
Kidd
Scianna
Colton
Lutheran
Allele
M
N
S
s
Fya
Fyb
K1
K2
Doa
Dob
Dia
Dib
Jka
JKb
Sc1
Sc2
Coa
Cob
Lua
Lub
Aua
Aub
Fragment length (bp)
162
300
239
239
167
167
145
145
182
182
336
336
244
244
155
155
191
191
173
173
146
146
Nucleotide mutation
71G/A
72T/G
143C/T
143C/T
131G/A
131G/A
578C/T
578C/T
793G/A
793G/A
2561C/T
2561C/T
838G/A
838G/A
169G/A
169G/A
134C/T
134C/T
230G/A
230G/A
1615A/G
1615A/G
Forward primer 5'-3'
CAGCATCAAGTACCACTGGT
TCAGCATTAAGTACCACTGAG
CGATGGACAAGTTGTCCCA
CGATGGACAAGTTGTCCCG
CAGCTGCTTCCAGGTTGGCAC
CAGCTGCTTCCAGGTTGGCAT
ACTCATCAGAAGTCTTTGCA
ACTCATCAGAAGTCTTTGCG
ATTCGATTTGGCCAATTCCTT
ATTCGATTTGGCCAATTCCTC
GGGCCAGGGAGGCCA
GGGCCAGGGAGGCCG
CCCAGAGTCCAAAGTAGATGTC
CCCAGAGTCCAAAGTAGATGTCT
CCTCCTTGGGTACCGTTTCC
CCTCCTTGGGTACCGTTTCT
GAACAACCAGACGGC
GAACAACCAGACGGT
CATCTCAGCCGAGGCTAAAAC
CATCTCAGCCGAGGCTAAAAT
CACCTCAGTCACTCACGCGC
CACCTCAGTCACTCACGCGT
Reverse primer 3'-5'
AGCTCGCATTTCTCAGTGTTTG
AGCTCGCATTTCTCAGTGTTTG
CATGTGGGTGGCACCCTGCC
CATGTGGGTGGCACCCTGCC
ATGTCCACAGTCACTCGCCA
ATGTCCACAGTCACTCGCCA
GCTCCCCCAGCCCCCGTCCG
GCTCCCCCAGCCCCCGTCCG
GTTTACCCGTTCTGCTAA
GTTTACCCGTTCTGCTAA
CCTGCCAGCTCCATGTGAC
CCTGCCAGCTCCATGTGAC
CAGGACGGACAAAGGA
CAGGACGGACAAAGGA
TCCTGTGGCAGCCTAAGAG
TCCTGTGGCAGCCTAAGAG
GTTTCTTGGAGCAGGTTAAACA
GTTTCTTGGAGCAGGTTAAACA
CTGCACTGTGAAGCTCTCAC
CTGCACTGTGAAGCTCTCAC
CTGCACTGTGAAGCTCTCCA
CTGCACTGTGAAGCTCTCCA
Electrophoresis
Each amplified product (5-10 mL) was loaded in a certain sequence onto a 2.5% agarose
gel in about 100 mL of 0.5X TBE buffer. Electrophoresis was conducted under 150 V and 110 mA
current for 12 min. The electrophoresis stopped when the internal control bands and the positive
bands were separated. The gel was stained with ethidium bromide, imaged, and analyzed.
Quality control
Each sample contained the internal control, which was a 429-bp conserved fragment
Genetics and Molecular Research 16 (1): gmr16019176
G.Y. Lin et al.
6
of HGH. This internal control was run in each experiment and could be detected during
electrophoresis. The amplified product of the internal control was also visible in the negative
wells. The internal control bands in the positive wells were very weak or non-existent likely
because the specific primers competed with internal control primers for substrates such as Taq
polymerase during amplification.
PCR-SSP method is a qualitative assay and has extremely high detection sensitivity.
Any improper operation will lead to false negative or false positive results. In addition to strict
quality control measures (e.g., internal control), the experiment was repeated if the results
were ambiguous for some alleles. All experimental results were reported and reviewed by two
professionals experienced in matching in kidney transplantation using the PCR-SSP method.
Result interpretation
Intensity of the specific bands did not interfere with result interpretation since PCRSSP is a qualitative assay. The typing results were interpreted according to the standard typing
result table (provided by Tianjin Super Biotechnology Development Co., Ltd. Tianjin city, in
China.). Criteria for interpretation were as follows: 1) positive wells: simultaneous appearance
of internal control bands and specific amplification bands was considered positive; 2) negative
cells: appearance of only internal control bands, but no specific amplification bands, was
considered negative; 3) appearance of neither internal control bands nor specific amplification
bands was regarded as a failure, and a second detection was needed.
Statistical analysis
The SPSS 12.0 software (In the biological network to provide biological software,
SPSS12.0 free version.) was used for statistical analysis. According to the Hardy-Weinberg
principle, for a sufficiently large population of sexual reproduction, the allele frequencies will
remain constant across generations if the individuals are allowed random breeding without
gene mutation, introduction of new genes, or natural selection. Therefore, the sum of allele
frequencies will be 1, and the sum of all genotype frequencies will be 1 or 100%. Now consider
a diploid individual who has two alleles, A and a, on one gene locus. The population has N
such individuals. The number of individuals carrying AA, Aa, and aa genotypes is n1, n2, and
n3, respectively. The following formulae were used for calculation: (1) frequency of A allele
= number of individuals carrying A allele/(number of individuals carrying A allele + number
of individuals carrying a allele) n1/N + n2/2N; (2) frequency of AA genotype = number of
individuals carrying AA genotype/total number of the diploid individuals in this population
= n1/N; (3) and relationship between allele frequency and genotype frequency: frequency of
allele A = frequency of AA genotype + 1/2 frequency of Aa genotype.
In a natural population, the genotype frequencies are not calculated based on allele
frequencies unless the relationship between the two obeys the Hardy-Weinberg equilibrium.
The calculation formulae are as follows: 1) frequency of homozygous genotype is the square
of the allele frequency, i.e., AA = A2; 2) frequency of heterozygous genotype is the product
of frequencies of two alleles multiplied by 2, i.e., Aa = Ax a x 2; 3) expected value of the
frequency of homozygous genotype = total number of individuals x the square of the frequency
of homozygous genotype; 4) expected value of the frequency of heterozygous genotype =
the total number of individuals x the frequency of homozygous genotype x the frequency of
Genetics and Molecular Research 16 (1): gmr16019176
7
Rare blood type genes in the Uygur population
heterozygous genotype x 2; and 5) the expected and observed values of genotype frequencies
were compared using the chi-square test to determine whether the Hardy-Weinberg equilibrium
was reached. The calculation formula was chi-square (c2) = (observed value - expected value)2/
expected value. P < 0.05 was considered to indicate a significant difference. The statistical
processes were implemented by experienced researchers.
RESULTS
Genotype frequencies of the 9 rare blood groups were in Hardy-Weinberg equilibrium
in Uygur population
The phenotype frequencies of the 9 rare blood groups obeyed the Hardy-Weinberg
equilibrium in Uygur population, without significant difference in the expected and observed
values of frequencies (P > 0.05). The allele frequencies remained stable. Rare phenotypes were
found in 2 rare blood groups, which were S-s- phenotype in 4 cases with a frequency of 0.0253
and Jk(a-b-) phenotype in 1 case with a frequency of 0.0063. This has not been previously
reported for the Uygur population. Monomorphism was found in the allele frequencies of
Scianna, Colton, and Lutheran blood groups. A skewed distribution was apparent for the allele
frequencies of Kell and Diego blood groups. Table 2 shows the phenotype frequencies of the
other 4 rare blood groups.
Table 2. Allele frequencies of four rare blood groups and their Hardy-Weinberg equilibrium status in the Uygur
population of Xinjiang.
Rare blood group
MNS
Duffy
Kell
Dombrock
Diego
Kidd
Scianna
Colton
Lutheran
Phenotype
M+NM-N+
M+N+
S+sS-s+
S+s+
S-sFya+bFya-b+
Fya+b+
K1+2K1-2+
K1+2+
Doa+bDoa-b+
Doa+b+
Dia+bDia-b+
Dia+b+
Jka+bJka-b+
Jka+b+
Jka-bSc1+2Sc1-2+
Coa+bCoa-b+
Coa+b+
Lua+bLua-b+
Au a+bAu a-b+
Aua+b+
Observed value (frequency)
53 (0.3354)
28 (0.1772)
77 (0.4873)
6 (0.0380)
105 (0.6646)
43 (0.2725)
4 (0.0253)
81 (0.5127)
18 (0.1139)
59 (0.3734)
0 (0)
153 (0.9684)
5 (0.03165)
9 (0.0570)
93 (0.5886)
56 (0.3544)
0 (0)
143 (0.9051)
15 (0.0949)
45 (0.2848)
31 (0.1962)
81 (0.5127)
1 (0.0063)
158 (1.0000)
0 (0)
157 (0.9937)
0 (0)
1 (0.0063)
0 (0)
158 (1.0000)
99 (0.6266)
1 (0.0063)
58 (0.3671)
Expected value (frequency)
52.99 (0.3353)
27.99 (0.1772)
77.02 (0.4873)
4.80 (0.0304)
101.35 (0.6415)
44.11 (0.2792)
7.74 (0.0489)
77.29 (0.4892)
14.28 (0.0904)
66.44 (0.4205)
0.04 (0.0002)
153.00 (0.9684)
4.91 (0.0311)
8.67 (0.0549)
92.66 (0.5865)
56.67 (0.3587)
0.36 (0.0063)
143.38 (0.9075)
14.27 (0.0903)
46.28 (0.2929)
32.37 (0.2049)
77.40 (0.4899)
158.00 (1.0000)
0 (0)
156.11 (0.9880)
0.01 (0)
1.85 (0.0001)
0 (0)
158 (1.0000)
103.71 (0.6564)
5.70 (0.0361)
48.62 (0.3077)
Genetics and Molecular Research 16 (1): gmr16019176
Allele frequency
0.5791
0.4209
0.1743
0.8009
0.6994
0.3006
0.0158
0.9842
0.2342
0.7658
0.0474
0.9526
0.5412
0.4526
0.167
1.0000
0
0.9940
0.0059
0.391
0
1.0000
0.8102
0.1899
1.810
2
0
0
0
0.300
0.131
0
1.807
0.178
0.969
0.833
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0.214
3.86
P
>0.05
>0.05
>0.05
>0.05
>0.05
>0.05
>0.05
>0.05
>0.05
>0.05
>0.05
8
G.Y. Lin et al.
Comparison of genotype and allele frequencies of 9 rare blood groups between the
Uygur population and other ethnic minorities
The allele frequencies are shown in Table 3. Allele frequencies of 9 rare blood groups
were obtained among the Uygur population, which were M = 0.5791, N = 0.4209, S = 0.1743,
s = 0.8009, Fya = 0.6994, Fyb = 0.3006, K1 = 0.0158, K2 = 0.9842, Doa = 0.2342, Dob =
0.7658, Dia = 0.0474, Dib = 0.9526, JKa = 0.5412, JKb = 0.4526, Sc1 = 1.0000, Sc2 = 0, Coa
= 0.9940, Cob = 0.0059, Lua = 0, Lub = 1.0000, Aua = 0.8102, and Aub = 0.1899, respectively.
Polymorphism was found in the allele frequencies of 5 rare blood groups, which were MNS,
Kidd, Duffy, Dombrock, and Diego. The allele frequencies of the 9 rare blood groups displayed
unique features among the Uygur population. Table 3 shows the comparison of the allele
frequencies of the 9 rare blood groups among different ethnic minorities in China.
Table 3. Comparison of the allele frequencies of the 9 rare blood groups among different ethnic minorities in China.
Rare
blood groups
MNS
Duffy
Kell
Dombrock
Diego
Kidd
Scianna
Colton
Lutheran
Allele
M
N
S
s
Fya
Fyb
K1
K2
Doa
Dob
Dia
Dib
JKa
JKb
Sc1
Sc2
Coa
Cob
Lua
Lub
Aua
Aub
Frequency
Uygur population
of Xinjiang
(N = 158)
0.5971
0.4209
0.1743
0.8009
0.6994
0.3006
0.0158
0.9842
0.2342
0.7658
0.0474
0.9526
0.5412
0.4526
1.0000
0
0.9940
0.0059
0
1.0000
0.8102
0.1899
Frequency
Kazak population
of Xinjiang
(N = 196)
0.645 4
0.354 6
0.142 9
0.846 9
0.757 6
0.242 3
0.002 6
0.997 5
0.221 9
0.778 0
0.035 7
0.964 3
0.538 3
0.456 7
1.000 0
0
0.992 4
0.007 7
0
1.000 0
0.849 5
0.150 5
Frequency
Hui population
of Xinjiang
(N = 220)
0.5204
0.4796
0.0954
0.9046
0.8727
0.1273
0.0068
0.9932
0.1386
0.8614
0.0159
0.9841
0.4953
0.5045
1.0000
0
0.9932
0.0068
0
1.0000
0.9091
0.0909
Frequency
Tibetan population
of Tibet
(N = 409)
0.6809
0.3191
0.1467
0.8533
0.9218
0.0782
0
1.000
0.1504
0.8496
0.0342
0.9658
0.5513
0.4487
1.0000
0
0.9976
0.0024
0
1.000 0
0.9108
0.0892
Frequency
Yi population
of Sichuan
(N = 240)
0.6833
0.3167
0.9170
0.9083
0.9458
0.0292
0
1.0000
0.1250
0.8750
0.0167
0.9833
0.5250
0.4750
1.0000
0
1.0000
0
0
1.0000
1.0000
0
Frequency
Han population
of Chengdu
(N = 332)
0.5752
0.4246
0.0376
0.9622
0.0663
0.9337
0.0407
0.9593
0.4428
0.5572
-
Electrophoresis results of genotypes of 9 rare blood groups in one Uygur subject
The electrophoretograms of the genotypes at 22 gene loci are presented in Figure 1.
The bands were clear and the positive bands were distinguished from the internal control bands
with parallel and equal displacement. The results can be easily interpreted.
DISCUSSION
The frequency of the M allele in the MN blood group system was detected as 0.5791
in the Uygur population of Xinjiang. It is consistent with the frequency of the M allele in the
Han population of Tai’an City in Shandong Province, the Han population of Jiangxi, the Han
population in Guangdong and Fujian Province, and the Han population in Chengdu of Sichuan
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Rare blood type genes in the Uygur population
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Figure 1. Electrophoretograms of genotypes of 9 rare blood groups in one Uygur subject. Genotypes of the MNS,
Duffy, Kell, Dombrock, Diego, Kidd, Scianna, Coltonand Lutheran rare blood groups were M+N+, S-s+, Fya+b+, K1-2+,
Doa-b+, Dia-b+, Jka+b+, Sc1+2-, Coa+b-, Lua-b+, and Aua-b+, respectively.
province, which is 0.5229, 0.5428, 0.5827, and 0.5752, respectively (Meng et al., 2001; Li et
al., 2001; Ye et al., 2001; Guo and Guo, 2005; Hong et al., 2012; Zhang et al., 2014). However,
the frequency of the M allele in the Uygur population is lower than that found in the Tibetan
population in Tibet, which is 0.6809 (Zhang et al., 2014). The frequency of N allele was 0.4209
in the Uygur population, which is also consistent with that found in other ethnic minorities of
China. The frequencies of S and s alleles are rarely reported in China. We found the frequency
of S allele in the Uygur population was 0.1743, which is much higher than that in the Tibetan
population of Tibet (0.1467) (Zhang et al., 2014), the Han population of Chengdu in Sichuan
Province (0.0376) (Hong et al., 2012), and the Hui population of Xinjiang (0.0954) (Lin et
al., 2016). Thus, the Uygur population displayed some unique features in allele frequencies
of the 9 rare blood groups. Moreover, there were 6, 105, and 43 cases carrying SS, ss, and Ss
genotypes, respectively, and a rare S-s- phenotype was found in 2 cases, with a frequency of
only 0.0102. The S-s- phenotype has not been reported previously in China. The alleles of the
9 rare blood groups in the Uygur population of Xinjiang are featured by high expression and
high polymorphism. Based on this feature, countermeasures can be adopted for meeting the
blood transfusion needs in ethnic minorities in Western China.
The Duffy blood group system was first discovered in 1950 (Zhao and Li, 2009). As an
important erythrocyte blood group, the Duffy blood group mainly produces the IgG1 antibody,
which can cause acute and delayed transfusion reactions. The frequency of Fya antigens is
above 0.9700 in China, Japan, and North Korea; not above 0.6600 in individuals of Europeandescent, 0.1000 in Africans, and 0 in individuals belonging to the Central African Republic,
and all the Central African individuals carry the Fy(a-b-) phenotype. However, the Fy (a-b-)
phenotype is very rare in individuals of European-descent and Asians (Zhao and Li, 2009).
Genetics and Molecular Research 16 (1): gmr16019176
G.Y. Lin et al.
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In our study, the frequency of Fya antigen was 0.6694 in the Uygur population of Xinjiang,
which is significantly lower than that found in the Tibetan population of Tibet (0.9218) (Zhang
et al., 2014), the Han population of Jiangsu (0.9400) (Liu et al., 2012), the Han population
of Luoyang (0.9527) (Yang et al., 2012), the Han population of Zhejiang (0.9412) (Fu et al.,
2001a) and the Han population of Shanghai (0.9649) (Zhao and Li, 2009). It is easy to see that
given the unique allele frequencies of the rare blood groups in Uygur population, more efforts
should be devoted to the screening of rare blood groups in case of emergency use.
In the Dombrock blood group system, the frequency of the Doa antigen is lower in
individuals of African-descent, American Indians, and Asians, and lower in individuals of
European-descent belonging to Northern Europe and USA. The positive rate of Doa antigen
is 67% in individuals of European-descent, 55% in individuals of African-descent, 24% in
the Japanese, and 0.1400 in Thai people (Delanghe et al., 1995; Grassineau et al., 2007; Li et
al., 2010). The frequency of the Doa antigen was 0.2342 in the Uygur population of Xinjiang,
which is consistent with that reported for the Uygur population of Xinjiang (0.2559) (Wu et
al., 2001); however, it is much higher than that in the Tibetan population of Tibet and the Han
population of Chengdu in Sichuan Province (0.0663) (Hong et al., 2012), the Han population
of Shanghai (0.0614) (Zhao and Li, 2009), and the Han population of Xi’an City (0.1159) (Liu
and Liu, 2003). This fully demonstrates the large variation in allele frequencies of the rare
blood groups across the regions and ethnic groups.
The frequency of Dia antigen in the Diego blood group system is 0.0474 in the Uygur
population of Xinjiang, which is much higher than that found in the Tibetan population of Tibet
(0.0342) (Zhang et al., 2014), the Han population of Chengdu in Sichuan Province (0.0407)
(Hong et al., 2012), the Han population of Xi’an City (0.0250) (Liu and Liu, 2003), and the
Han population of Jiangxi Province (0.0198) (Xiao et al., 2010). Given the higher frequency
of Dia antigen in the Uygur population of Xinjiang, more efforts should be undertaken in
preparation for emergency blood transfusion for the Uygur population. However, in Zhang et
al. (2014), the allele frequencies of Duffy, Kell, Diego, Scianna, Colton, and Lutheran blood
groups in the Uygur population are reported as similar to those in other studies. We found that
the frequency of the Dia antigen in the Uygur population of Xinjiang was different compared
with other Chinese ethnic minorities.
The frequencies of antigens of the Kidd blood group vary from one ethnic group
to another. In individuals of European-descent and African-descent, the frequency of Jka
antigen is slightly higher than that of Jkb antigen (Hashmi et al., 2007). In the Chinese Han
population, the frequency of the Jkb antigen is slightly higher than that of the Jka antigen (Fu et
al., 2001b; Song et al., 2008). The frequency of the Jka and Jkb antigens is 0.54212 and 0.4526,
respectively, in the Uygur population of Xinjiang. The allele distribution of the Kidd blood
group in the Uygur population is more similar to that of individuals of European-descent and
different from the Chinese Han population. The frequency of the Jka antigen (0.5421) in the
Uygur population, is consistent with that found in the Tibetan population of Tibet (0.5513)
(Zhang et al., 2014), the Uygur population of Xinjiang (0.5429) (Qiu et al., 2012), the Han
population of Zhejiang (0.4902) (Fu et al., 2001b), the Han population of Chengdu (0.4428)
(Hong et al., 2012), the Hui population of Xinjiang (0.4504) (Qiu et al., 2012), and the Hui
population of Xinjiang in another studies (0.4955) (Lin et al., 2016). Apparently, the Uygur
population of Xinjiang exhibits a distinctive feature in the allele frequencies of the rare blood
groups as compared with the Han and Hui population. The reasons mainly lie in the origin and
migration of the ethnic group.
Genetics and Molecular Research 16 (1): gmr16019176
Rare blood type genes in the Uygur population
11
It is reported that the frequency of the Jk(a-b-) phenotype is below 0.01% in the Asian
population but higher in Indians, Bolivians, and individuals of European-descent (Song et
al., 2008). This phenotype has been found in Macao Blood Center (Liu, 2011). Individuals
carrying this rare phenotype are difficult to identify using serological tests (Li et al., 2010).
Yan (2007) identified 10 individuals out of 50,034 donors carrying the Jk(a-b-) phenotype,
with a frequency of 0.0002. We reported a frequency of 0.0272 in the Hui population of
Xinjiang (Lin et al., 2016), with 6 subjects carrying this rare phenotype. This phenotype has
not been yet discovered in the Tibetan population of Tibet (Zhang et al., 2014). This study
identified a single Uygur subject carrying the Jk(a-b-) phenotype, with a frequency of 0.0051,
which is higher than that among the Asian population (0.01%). Therefore, it is demonstrated
that genetic migration among the Uygur population of Xinjiang is related to individuals of
European-descent in middle Asia, and this finding is of high significance for blood transfusion
and construction of the rare blood group library for ethnic minorities. More work should be
done in rare blood group screening based on a full exploitation of the local rare blood group
resource.
The observed and expected values of allele and genotype frequencies of 9 rare blood
groups were compared using the chi-square test, and no significant difference was found (P
> 0.05). The frequencies obeyed the Hardy-Weinberg equilibrium, indicating constant allele
frequencies of rare blood groups in Uygur population of Xinjiang.
A skewed distribution pattern was found for the allele frequencies of 2 rare blood
groups, Kell and Diego, for which the genotype frequency was 0. This agrees with the
distribution patterns reported for the Tibetan and Han populations. The allele frequencies of the
Scianna, Colton, and Lutheran rare blood groups indicated monomorphism, as was consistent
with the frequency distribution in the Tibetan and Han populations. The allele frequencies of
the MNS, Duffy, Kidd, and Dombrock rare blood groups indicated high polymorphism, which
is different from other ethnic minorities. In the comparison of the rare blood groups MNS,
Duffy, Kell, Dombrock, Diego and Kidd of Xinjiang Uygur, Kazak, Hui, Tibetan, Sichuan
and Chengdu, the highest frequency of gene was found in Xinjiang Uygur K1 0.0158, Doa
0.2342, The highest tribe of the Tibetan Han nationality is 0.6822, Dob 0.9337 and JKb 0.5572
are the highest; the Xinjiang Kazuya is the highest; the Tibetan Tibetan is the highest; the
Tibetan Tibetan Fyb 0.0782, JKa 0.5513 is the highest; the Tibetan Yi M 0.6833, S 0.9170 is
the highest; Gene antigen frequency were not the highest.The above results not only contribute
in understanding the genetic background of rare blood groups and historical migration for the
Uygur population of Xinjiang, but are also valuable for precision blood transfusion, reducing
hemolytic transfusion reaction, and matching in organ transplantation.
However, the sample size is limited due to limited funding. Moreover, the PCRSSP method is a qualitative and sensitive assay and some subjectivity is involved in result
interpretation, which could lead to errors.
Conflicts of interest
The authors declare no conflict of interest.
ACKNOWLEDGMENTS
Research supported by “474” People’s Liberation Army Hospital, Urmqi, China,
Genetics and Molecular Research 16 (1): gmr16019176
G.Y. Lin et al.
12
and compiled by the PCR Room Department of Blood Transfusion. Fund Project: Lanzhou
Military Region Medical and Health Research Project (#CLZ15JB26).
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