Đa dạng di truyền các giống bưởi ở đồng bằng sông cửu long dựa trên trình tự adn mã vạch và dấu phân tử issr

DA DANG DITRUYEN CAC GIONG Bl/OfI 6 DONG BANG SONG CUTl LONG
DUA TREN TRINH TU ADN MA VACH VA DAU PHAN TEf ISSR
Do Ian Khang1, Tram Thi Thanh Tien1, Tran Gia Huy1,
Nguyen Van Ay*2, Tran Thanh Men3

T6M TAT
Cac giong bddi d Dong bang song Cdu Long dddc khao sat trinh td ADN ma vach vdi 3 vbng trinh td ITS, ycflb,
psbK-psbl ket hop vdi phan tich da dang di truyen bang dau phan td ISSR. Ket qua khao sat cac viing trinh tv£ cho
thay cac giong biidi trong nghien cdu tUdng doi dong nhat vdi nhau ve mat di truyen qua phan tich cac vung trinh
tu ITS, ycflb, psbK-l. Ket qua PCR vdi moi ISSRK2 va ISSR22 da khuech dai dilqc 19 bang ADN trong do co 11 bang
da hinh chiem 57,89% va 8 bang don hinh chiem 42,11%. Dau phan td ISSRK2 phan biet diiqc biidi da xanh vdi
budi nam roi va budi ruby. Biidi dudng trang va budi thanh kieu co he so tUcfng dong den 95%. Nhu vay dua tren hai
dau phan td ISSRK2 va ISSR22 da cho thay sd da hinh trong trinh td cac giong bddi nghien cdu. Dieu nay cho thay
tiem nang cda dau phan td ISSR trong phan tich da dang di truyen cac giong bddi, phuc vu cho cong tac chon giong.
Td khoa. Bddi, da dang di truyen, ma vach ADN, dau phan td ISSR

! Vien Nghien cdu va Phat trien Cong nghe Sinh hoc, Trddng Dai hoc Can Tho
2Khoa Nong nghiep, Trddng Dai hoc Can Tho;3Khoa Khoa hoc Td nhien, Trddng Dai hoc Can Tho
19


Tap chf Khoa hoc va Cong nghe Nong nghiep Viit Nam - So 05(126)/202 7
i . d At v a n d e

Viet Nam thuoc khu vdc Dong Nam A, la mot
trong nhdng trung tam phat sinh cua cac loai cay
co mui (Rainer, 1975). Mot so cong trinh da cong
bo cho thay Viet Nam co nguon gen cay co mui kha
da dang vdi nhieu vung trong bddi truyen thong co
nhieu giong bddi quy nhd bddi dien, bddi thanh
tra, biidi da xanh, biidi bien hoa, biidi nam roi, ...

(Nguyen Thi Tuyet va ctv., 2018; Nguyen Phiiong
Thuy va ctv., 2016). Trong nam 2017, niidc ta xuat
khau hon 10.000 tan biidi, tang gap doi so vdi nam
2016 d hau het thi triidng ldn nhii: My, EU, Canada
va cac niidc Trung Dong (Tap chi Doanh nhan Viet
Nam, 2020). Ngay nay, de xac dinh va nhan dien
cac giong loai khac nhau hoac cac loai co dac diem
hinh thai tddng til nhau co rat nhieu phiiOng phap.
Phiiong phap dddc dung chu yeu pho bien dda tren
phan tich hinh thai cua mau vat. Tuy vay, phddng
phap hinh thai hoc thiidng gap nhieu trd ngai.
Trong boi canh hien tai d midc ta, cac nghien cdu
ve sd dung ADN ma vach de phan tich cac loai khac
nhau dang dan dddc quan tam va phat trien. Biidi
duac trong khap cac tinh dong bang song Cdu Long
(DBSCL), m6i vung deu co nhflng dieu kien sinh
thai nhat dinh anh hiidng den qua trinh song cung
nhii bao ton nguon gen. Viec nghien cdu da dang di
truyen nguon gen biidi da xanh d khu viic DBSCL
co y nghia trong viec bao ton tinh da dang sinh hoc
va sd dung co hieu qua cac nguon gen quy do phuc
vu cho cong tac chon tao giong. De tai diidc thiic
hien nham phan tich diidc da dang trinh td ADN

ma vach (ITS, ycflb, psbK-psbl) va danh gia da dang
di truyen cac giong bddi d nam tinh trong khu vdc
DBSCL bang dau phan td ISSR.
II. VAT LIEU VA PHUCfNG PHAP NGHIEN CUU
2.1. Vat lieu nghien cdu
Mau la bddi da xanh (Citrus maxima) dddc thu

tai 5 tinh gom: Tien Giang, Soc Trang, Hau Giang,
Ben Tre, Vinh Long. Bddi “Ruby”, bddi thanh kieu
(Thdi An - 6 Mon - Can Thd), bddi dddng trang,
bddi nam roi.
2.2. Phddng phap nghien cdu
2.2.1. Thu mdu va tach chietAD N
Tien hanh trich ADN td mau la, ADN dddc chiet
tach theo quy trinh cua Doyle va Doyle (1990). Kiem
tra chat lddng cua ADN ly trich dddc bang phddng
phap dien di tren gel agarose 1% d hieu dien the
100 V trong 30 phut. Ban gel dddc quan sat dddi anh
sang td ngoai tren he thong may Bio-Rad UV 2.000.
Nong do ADN dddc xac dinh bang phddng phap do
quang pho d bddc song 260 va 280 nm.
2.2.2. Khuech dai vung trinh tu ITS, ycflb, psbK-l
Moi phan dng PCR 50 pi gom co 25 pL nddc khd
ion; 20 pL master mix 2X (Buffer, MgCl2, dNTPs,
Taq polymerase); 1 pL moi xuoi va moi ngddc va
3 pL ADN khuon. Chu ky nhiet PCR khuech dai cua
3 vung trinh td ITS, ycflb va psbK-l dddc the hien
dbang 1.

Bang 1. Trinh td va chu ky nhiet cua cac moi sd dung trong nghien cdu
Trinh td

Chu ky nhiet

ycfl fo F: TCTCGACGAAAATCAGATTGTTGTGAAT
W ib-R ACATGTCAAGTGATGGAAAA (Dong et al, 2015)

94°C -5 phut; 94°C - 30 giay, 55°C - 40 giay,
72°C -1 phut (35 chu ky); 72°C -10 phut

psbK-I-F:TGCCTTTGTTTGGCAAG
psbK-I-RiAGAGTTTGAGAGAGCAT (Reginato etal, 2010)

94°C - 5 phut; 94°C - 30 giay, 55°C - 40 giay,
72°C - 40 giay (35 chu ky); 72°C - 5 phut

ITS1-F: TCCGGGAACCTGCGG
ITS4-R; TCCTCCGCTTTGATGC
(“White et al., 1990)

95°C - 5 phut; 95°C - 30 giay, 58°C - 30 giay,
72°C - 1 phut (35 chu ky); 72°C - 7 phut

ISSR22: TGTGTGTGTGTGTGTGCA
ISSRK2: GTGGTGGTGGTGAC (Samriti et al., 2017)

94°C - 4 phut; 94°C - 1 phut, 50°C - 45 giay,
72°C - 2 phut (35 chu ty); 72°C - 7 phut

2 2 1 Pkmm tkh so lieu
K£t qua giai trinh td ADN dddc kiem tra tren
d n a a t o p a m bang chddng trinh Bioedit. Td cac
«■»*» H c m cac giong bddi da xanh, xac dinh trinh
W d o n g (Consensus sequence) cho giong bddi nay
m a a n i l t Ialignment) vdi cac trinh td cua cac giong
riba kM e tnm g n^iien cdu de xac dinh cac

SNPs tiem nang. So sanh sd tddng dong gifla cac
giong bddi trong nghien cdu vdi co sd dd lieu cua
ngan hang gene NCBI.
San pham khuech dai dau phan td ISSR dddc
phan tich bang phan mem NTSYSpc2. Bieu do hinh
cay ve moi quan he phat sinh loai dddc xay ddng
theo phddng phap UPGMA.


Tap chi Khoa hoc va Cong nghe Nong nghiep Viet Nam - So 05(126)/2021

m . KET QUA VA THAO LUAN

ro ret. Khi so sanh vdi thang chuan thitong mai co
kich thudc 3 kb cho ket qua khuech dai dupe chieu
dai cac doan gen lan lupt la 900 bp, 500 bp, 700 bp
tUdng ling vdi 3 moi la ycflb (Hinh 1A), psbK-psbl
(Hinh IB) va ITS (Hinh 1C). Tif nhQng ket qua dat
dupe cho thay san pham PCR tren da du dieu kien
de tien hanh budc giai trinh tU.

3.1. Khuech dai va giai trin h tit
Ket qua dien di san pham PCR khuech dai vimg
trinh tU ITS, ycfl b va psbK-psbl dupe thilc hien tren
gel agarose 2%, sau khi kiem tra ket qua cho thay
vdi 3 moi tren dieu cho hien cac bang ADN sang

r
1500bp---800bp—

2 3

4

5 6 7

8

9 10

500bp

1 2

3

4

5

6

7

8

9

10

11

700bp-----

C

Hinh 1. Pho dien di san pham khuech dai geneycflb (A), trinh tilpsbK-1 (B)
va ITS (C) cua cac giong budi trong nghien ciiu
Ghi chu: (1) thangchuan 3kb; (2) biidi daxanh Tiengiang(BDTG) + doi chvingdudng; (3) budi daxanh Soc Trdng
(ST);(4) budi da xanh Hau Giang (BDHG); (5) btidi da xanh Vinh Long (BVL); (6) budi da xanh Ben Tre (BDB); (7)
budi ruby (BRB1); (8) budi thanh kieu (BTK); (9) budi dudng trdng (BDT); (10) budi 5 roi (STR); (11) Doi chUng dm.
Cac doan ADN ma vach cua 9 giong biidi tren
vung trinh til ycfl b sau khi giai trinh til co chieu dai
800 - 820 bp, ket qua dung nhii dil kien ban dau la
khoang 800 - 900 bp. Cac doan ADN ma vach cua
9 giong biidi tren moi psbK-psbl sau khi giai trinh
til co chieu dai 430 - 440 bp, ket qua cung dat dung
nhil dil kien ban dau khi gbi trinh til la khoang
400 - 500 bp. Cac doan ADN ma vach cua cac giong
biidi tren vimg trinh til ITS sau khi giai trinh til co
chieu dai khoang 600 - 700 bp. Ket qua giai trinh til
cho thay cac doan trinh til tren vimg trinh til ITS co
chieu dai san pham khuech dai xap xi vdi nghien ciiu
cua Kyndt va cong tac vien (2010) vimg ITS trong
nghien ciiu tren co chieu dai la 703, cac trinh til thu
dupe tot, khong bi nhieu, du co sd de phan tich.
3.2. Phan tich trinh tU ADN
Ket qua phan tich vung trinh til ycflb cua cac
giong budi trong nghien ciiu cho thay khong co sil
khac biet qua ldn trong ciing mot giong cung nhu

gifla cac giong budi khac nhau cua cac doi tupng
dupe nghien ciiu. Qua do cho thay gene ycfl b co

tinh bao ton cao giQa cac giong budi thu dupe trong
khu vile DBSCL. Khi phan tich vung trinh tU ITS
cho thay mot so vi tri sai khac d giong budi ruby va
dot bien diem mat 1 nucleotide C d giong budi ruby,
budi thanh kieu va 1 nucleotide C d budi dudng
trang. Co 2 vi tri xuat hien dot bien mat 1 nucleotide
d vi tri 10 hien dien d giong budi ruby, budi thanh
kieu va vi tri thd 21 d giong budi dudng trang. Ben
canh do, nghien ciiu cung xac dinh dupe 3 SNPs cua
giong ruby so vdi cac giong con lai.
Cac trinh tU gen ycfl b va psbK-psbl cua cac giong
budi khong co nhieu tren ngan hang gen do day la
hai trinh tU mdi, chUa dupe nghien ciiu nhieu. Chi
Citrus chi co 1 nghien ciiu cua Xu va cong tac vien
(2016) tai Trung Quoc. Nghien ciiu nay da giai ma
toan bo genome trong he luc lap cua budi da xanh.
Vi vay, trinh tU nay dupe sh dung de lam cP sd so
sanh, doi chieu vdi cac trinh til trong nghien ciiu
nay. Rieng vung ITS la trinh tU pho bien nen dupe
tim thay nhieu tren cP sd dQ lieu.
21


Tap chi Khoa hoc va Cong nghe Nong nghiep Viet Nam - So 05(126)/2021

Theo Dong va cong tac vien (2015), ycflb dddc
xem nhd dau phan td co kha nang nhan dien cao hon

ca matK va rbcL day la 2 vung trinh til pho bien dddc
s£i dung rang rai de lam dau phan td, ycfl b dddc xem
la dau phan td hieu qua trong nhan dien phan td d
cac cap do phan loai thap. Tuy nhien, trong nghien
cdu dddc thdc hien d cap do cao hdn tren cay co mui
ycflb lai khong the hien sd da hinh trong trinh td.
Td do cho thay rang viec sd dung ycfl b de phan loai
d thdc vat noi chung va cay co mui noi rieng khong
that sd hieu qua.
d vung trinh td ITS co xuat hien nhieu sai khac
tuy chiia the ket luan no co phai la ma vach tren cay
co mui hay khong, nhdng day cung cho thay dddc sd
da dang di truyen tren vimg trinh td ITS d bddi do vi
tri sai khac co the xuat hien trong cung giong hoac
khac giong. Dieu nay cung phu hop vdi nghien cdu

0.66

0.73

cua Tran Thanh Men va cong tac vien (2008) tien
hanh nghien cdu da dang sinh hoc cac giong bddi d
Viet Nam.
3.3. Moi quan he di truyen gifia cac giong bddi
bang dau phan td ISSR
Ket qua dien di dau phan td ISSRK2 cho xuat
hien 11 bang trong do co 8 bang da hinh chiem ty le
72,73% so vdi tong so. Ket qua dien di dau phan td
ISSR22 cho xuat hien 8 bang trong do co 3 bang da
hinh chiem ty le 37,5% so vdi tong so.

Theo ket qua d hinh 2 vdi do tddng dong dao
dong trong khoang 0,66 - 0,95 vdi khoang dao dong
la 0,07 ddpc chia thanh 3 nhom chinh: nhom I vdi
Nhom III co he so tddng dong la 0,76, nhom I, II vdi
nhom III co he so tiidng dong thap nhat la 0,66.

0.81
C oefficient

0.88

0.95

Hinh 2. Gian do pha he the hien moi tUdng quan di truyen cua 9 giong biidi
Nhom I: Chi co duy nhat mau budi da xanh Tien
Giang (BDTG). Nhom II: trong nhom nay lai dddc
chia lam 3 nhom nho la: (II.l) gom biidi da xanh Soc
Trang (ST) va bddi da xanh Hau Giang (BDHG) vdi
he so tddng dong la 0,89. II.2: Gom bddi da xanh
VInh Long (BVL) va bddi da xanh Ben Tre (BDB)
wji he so tddng dong la 0,94. II.3: Gom bddi thanh
kieu (BTK) va bddi dddng trang (BDT) vdi he so
tifdng dong gan 0,95. Nhom III: chi co 2 giong la
bddi ruby (BRB) va bddi nam roi (STR) vdi he so
U n f dong la 0,74. Tuy nhien ket qua tren cho thay
trong cung mot giong bddi da xanh lai d 2 nhom
k k k nhan, la giong tai Tien Giang (BDTG) co khac
I s x U n wdi he so tddng dong la 0,76 vdi cac giong
bddi da xanh d cac tinh con lai trong nghien cdu
A Sdc Trang (ST), Hau Giang (BDHG), Vinh Long

(PITJL B£b The (BDB), trong khi 4 mau bddi da
iH fe nay Ip cung nam trong nhom II vdi bddi thanh
Kiiu litXlk]) xa bddi dddng trang (BDT) nhdng van

co tach biet vdi 4 m lu bddi da xanh vdi he s6 tddng
dong la 0,85. Co nhieu yeu to tao nen sd khac biet
trong quan the nhd kha nang phat tan cua loai va sd
anh hddng cua moi trddng song (Kerdelhue et al,
2002) hoac do tac dong cua con ngddi. Quan diem
nay cung phu hdp vdi nghien cdu cua Snkiewicz
va cong tac vien (2001) khi nghien cdu ve moi lien
he di truyen cua cac giong cay trong trong canh tac
nong nghiep cho thay con ngddi la yeu to quan trong
trong viec di chuyen giong cay trong gida cac vung
dia ly. Tuy nhien, ket qua nay chi la tam thdi dda tren
hai dau phan td trong nghien cdu. De tang cao ket
qua can phan tich them nhieu dau phan td hdn nda.
Dda vao hinh 3 cho thay, vi tri bang phan biet
bddi da xanh vdi bddi 5 roi va bddi “Ruby” d vi tri
khoang 450 bp (mui ten mau trang). Cac mau bddi
da danh deu co bang nay trong khi bddi nam roi va
bddi “Ruby” khong co.


Tap chi' Khoa hoc va Cong nghe Nong nghiep Viet Nam - So 05(126)/2021

n

3000bp
1500bp

lOOObp
500bp -

SB

\

\\

Hinh 3. Ket qua dien di cac mau bddi vdi dau phan td ISSRK2
Ghi chu: gieng 1: thang chuan 100 bp (Omega Bio-tek); gieng2: Doi chiing dUdng; gieng 3: bildi da xanh Soc Trang;
gieng 4: btidi da xanh Hau Giang; gieng 5: bUdi da xanh Vinh Long; gieng 6: budi da xanh Ben Tre; gieng 7: budi ruby;
gieng 8: budi thanh kieu; gieng 9: budi diidng trang; gieng 10: budi nam roi.
Khuat Hutu Trung va cong tac vien (2009) da tien
hanh nghien cdu da dang di truyen tren 41 giong
btidi ban dia Viet Nam bang chi thi SSR lad cho ket
qua bddi da xanh nam ciing nhom vdi bddi nam roi
vdi he so tiicfng dong la 0,5 trong khi nghien cutu
dtipc thdc hien lai cho ket qua btidi da xanh va biidi
5 roi thupc 2 nhom khac nhau. Trong nghien cdu
nay, vdi dau phan td ISSR thi cho do tdcfng dong
0,66. N hd vay, co the thay cac chi thi phan td khac
nhau se cho ra do tdong dong khac nhau.
Trong nghien cdu nay co 2 giong la Bddi Thanh
Kieu (BTK) va bddi dddng trang (BDT) co do tdong
dong di truyen den 95% mac du rat khac biet ve
hinh thai. Do nddc ta co nhieu giong bddi co hinh
thai trai kha giong nhau nen de dda den nham lan
ten giuta cac giong bddi d nhutng vung khac nhau
(Vu Cong Hau, 2000).

IV. KET LUAN
Gen ycflb va trinh td psbK-psbl chda that sd la
mot trinh td ma vach tiem nang de phan loai cac
giong bddi trong nghien cdu. Khong co sd khac
biet trinh td d trong cung giong cung nhd d nhdng
giong khac nhau. Nhd vay cac viing trinh td ADN
ma vach trong nghien cdu chda phan biet ddpc cac
giong bddi, do do can chon cac trinh td ADN ma
vach khac de tiep tuc khao sat. Dau phan td ISSR co
the phan biet ddpc cac giong bddi do co sd da hinh,
nhdng can nhieu dau phan td hdn nda de sd phan
biet ddpc ro rang hon gida cac giong. Nghien cdu
tren se gop phan xay ddng nen tang cho cac nghien
cdu tiep theo tren chi Citrus cung nhd d thdc vat ve
dng dung ADN ma vach trong dinh danh.

TAI LIEU KHAM KHAO
Vii Cong Hau, 2000. Trong cay an qua d Viet Nam. Tai
ban lan 3. NXB Nong Nghiep TR Ho Chi Minh:
100-116.
Tran Thanh Men, Nguyen Thi Pha, Tran Nhan Ddng,
Ha Thanh Toan, Tina Kyndt, Godelieve Gheysen
va Marcelle Holsters, 2008. Nghien cdu da dang
sinh hoc cac giong bddi (Citrus maxima (Burm)
Merr.) d Viet Nam bang phddng phap PCR-RFLP.
Hoi nghi khoa hoc “cay an trai quan trong d DBSCL
14-20.
Khuat Hflu Trung, Ha Trong Huy, Nguyen Trddng
Khoa, Ngo Hong Binh, Nguyen Thanh Binh, Dang
Trong Ldofng va Le Huy Ham, 2009. Nghien cdu da

dang di truyen mot so giong bddi ban dia Viet Nam
(Citrus grandis) bang chi thi microsatellite. Tap chi
Cong nghe Sinh hoc, 7(4): 485-492.
Nguyen Phddng Thuy, Nguyen Thi Nga, Le Thi Cam
Tu, Dao Thi Be Bay va Tran Tlii Oanh Yen, 2016.
Danh gia dac tinh di truyen cua cac ddng bddi tif
nuoi cay hat nhd bang chi thi phan td microsatellites.
Tap chi Khoa hoc vd Cong nghe Viet Nam, 8: 30-35.
Tap chi Doanh nhan Viet Nam, 2020. Tim co hoi
cho trai bddi Viet Nam md rang ra thi trddng ldn,
o-rong-thi-truongcho-trai-buoi-viet-nam-22572.html, ngay truy cap:
20/5/2021.
Nguyen Thi Tuyet, Nguyln Thi Xuyen, Nguyen Thi
Lan Hoa, Bui Thi Thu Giang va Tran Danh Situ,
2018. Danh gia da dang di truy£'n mot so nguon gen
bddi (Citrus spp.) bang chi thi SSR. Tap chi Khoa hoc
Cong nghe Nong nghiep Viet Nam, 1(86): 14-18.
Dong, W.P., Xu, C., Li, C., Sun, J., Zuo, Y., Shi,
S., Cheng O., Guo J. and Zhou S., 2015. ycfl, the
most promising plastid DNA barcode of land planted
Scientific reports, 5(1): 1-5.
23


Tap chi Khoa hoc va Cong nghe Nong nghiep Viet Nam - So 05(126)/2021

Doyle, J.J. and Doyle, J.L., 1990. Isolation of plant DNA
from fresh tissue. Focus, 12:13-15.
Kerdelhue, C., Roux, G., Forichon, ]., Chambon, J.,
Robert, A. and Lieutier, F., 2002. (Curculionidae:

Scolytinae) on different pine species and validation
of T. destruens (woll.). Molecular Ecology, (11):
483-494.
Kyndt, T., Dung, T.N., Goetghebeur, R, Toan, H.T.
and Gheysen, G., 2010. Analysis of ITS of the
rDNA to infer phylogenetic relationships among
Vietnamese Citrus accessions. Genetic resources and
crop evolution, 57(2): 183-192.

Rainer, W. S., 1975. On the history and origin of Citrus.
Bulletin of the Torrey Bonical Club, 102(6): 369-375.
Snkiewicz, M., Gadamski, G. and Gawronski,
S.W., 2001. Genetic variation and phylogenetic
relationships of triazine resisnt and triazine
susceptible biotypes of Solanum nigrum analysis
using RAPD markers. Weed Res., 41: 287-300.
Xu, S-R., Huang, C-Y., Deng, Y-T., Zhou, L-L., Pan,
D-M., and Pan, H-L., 2016. The complete chloroplast
genome sequence of Citrus maxima (Burm.) Merr.
‘Guanximiyou’. Mitochondrial DNA, 5(1): 482-483.

Genetic diversity o f pomelo varieties in the Mekong Delta based
on DNA barcode and ISSR markers
Do Tan Khang, Tram Thi Thanh Tien, Tran Gia Huy,
Nguyen Van Ay, Tran Thanh Men
Abstract
Pomelo varieties in the Mekong Delta were examined by sequencing of three DNA barcode regions, including ITS,
ycflb, psbK-psbl in combination with genetic diversity analysis by ISSR markers. The results indicated that the
pomelo varieties in the study were similar in term of genetic diversity based on analyzing the sequences of ITS, ycflb
and psbK. The two ISSRK2 and ISSRK22 markers had amplified 19 DNA bands, including 11 polymorphic bands

accounting for 57.89% and 8 monomorphic bands for 42.11%, respectively. The marker ISSRK2 could distinguish Da
xanh from Nam Roi and Ruby pomelo varieties. Genetic similarity between Duong Trang and Thanh Kieu pomelo
varieties was 0.95. Therefore, based on the ISSR markers K2 and K22 the polymorphisms of pomelo varieties were
observed. The finding showed the potential of ISSR markers in analyzing genetic diversity of pomelo and could be
used in pomelo breeding.
Keywords: Pomelo variety, genetic diversity, DNA barcode, ISSR marker
Ngay nhan bai: 07/5/2021
Ngay phan bien: 18/5/2021

Ngiicfi phan bien: TS. Tran Ngoc Hung
Ngay duyet dang: 04/6/2021