全 文 ：广 西 植 物 Guihaia 30(3)：395～ 402 2O10年 5月
荞麦属植物淀粉酶和甲酸脱氢酶同功酶研究
张以忠1，2，陈庆富
(1．贵州师范大学 生命科学学院 植物遗传育种研究所，贵阳 550001；2．毕节学院 环境与生命科学系，贵州 毕节 551700)
摘 要：以聚丙烯酰胺凝胶电泳方法研究了荞麦属植物 8个种 42个收集系干种子和发芽种子的淀粉酶和甲酸
脱氢酶同功酶。结果表明，荞麦淀粉酶在干种子中缺乏活性，但是在发芽种子 中活性很强。在供试材料的发芽
种子中共发现 23个淀粉酶谱带，其 中甜荞和苦荞分别有 1O条和 8条。不同荞麦种间淀粉酶谱带差异很大，但
是同种内不同收集系问差异较小。谱带聚类分析表明大野荞和毛野养分别与甜荞和苦荞较近缘，支持它们分别
为甜荞和苦荞祖先种的假说 在干种子和发芽种子中，发现所有养麦种类均只有 1条位置一致的甲酸脱氢酶谱
带，暗示该酶在进化中具有高度稳定性。
关键词 ：普通荞麦 ；苦养 ；淀粉酶 ；甲酸脱氢酶 ；系统关系；起源与进化
CLC Number：$330 Document Code：A Article ID：1000—3142(2010)03—0395—08
Amylase and formate dehy
isozymes in the genus Fagopyrum
ZHANG Yi-Zhong ．CHEN Qing-Fu1
(1．Institute of Plant Genetics and Breeding，School of Life Science，Guizhou Normal University．Ouiyang
550001，China：2．Department of Environment and L fe Science，Bijie College，Bijie 551700，China)
Abstract：The amylase isozyme and formate dehydrogenase isozyme of 42 accessions of cultivated and wild buckwheat
belonging to eight species of the genus Fagopyrum including seven species of the big-achene group and one of the smal—
achene group were studied by means of polyacrylamide gel electrophoresis(PAGE)．The isozyme samples were obtained
from dry and sprouting seeds，respectively．The results showed that dry seeds had no amylase activity but the sprouting
seeds had very active amylase isozymes．A total of 23 bands for the amylase isozyme were observed in sprouting seeds of
cultivated and wild buckwheat，with F．esculentum and F．tataricum having 10 and 8 bands，respectively．Variations in the
amylase isozyme zymographs were large among different buckwheat species but were less so wi thin the same species．
The results of zymograph analysis and clustering showed that F．megaspartanimn and F．pilus are closely related to F．
esculentum and F．tataricum，respectively．supporting Chen’s hypothesis that F．megaspartanium and F．pilus are ances—
tors of these two species，respectively．Only one band was observed for the formate dehydrogermse isozyme in the dry and
spouting Seeds of al accessions．
Key words：common buckwheat；tartary buckwheat；amylase isozyme；formate dehydrogenase isozyme；phylogeny；
origin and evo[ution
Isozyme formation occurs when a living organism a—
dapts to its environment during the evolutionary process．
Isozymes show specificity in different species and gene—
ra，different tissues and organs，and during different de—
Reccived date：2008—10—15 Accepted date：2009—05—19
Foundation itern：Supported by the National Natural Science FOundation of China(3O270852，30471116)；Program for New Century Excellent Talents in
University(NCET-2004_0913)；Special P eet o{Development of Animal and Plant Varieties in Guizhou Pro~nce([2007]026)；Mega-projects of China
(2006BADO2H06)
Biography：ZHANG Yi-Zhong(1977一l。male．Master Degree。I ~turer．working with colege biological education and huekwheat research．
Author for corresp0ndence．E-mail：eqfl966@l63．eom
396 广 西 植 物 3O卷
velopmental stages(Hu & Wan，1985；Bao 8L Chen，
1993；A1ekhova et a1．，2001；Qu et a1．，2003；Liu et a1．，
2004；Yang et a1．，2006)．Using isozyme zymographs，it
is possible to determine gene existence，examine gene ex-
pression，and infer gene hereditary．At present，enzyme
isozymes are widely applied as genetic and biochemical
markers in studies of species identification，origin，evolu—
tion，and classification in many fields．A number of stud—
ies have shown that the amylase isozym e is a particularly
useful tool in phylogenetic studies and species identifica-
tion(Yuan ＆ Liang，1993；Wang etⅡ￡．，2000；Zhao et
a1．，2000；Jiang L Pen，2002；Hu eta1．，2003；Xie eta1．，
2004)．Formate dehydrogenase，on the other hand，al—
though it widely exists in higher plants，has scarcely
been reported．
Buckwheat，of which there are about 16 species
(Chen，1999a，b；Chen et a1．，2004)，belongs to the genus
Fagopyrum of the fami ly Polygonaceae(W u，1983；Lee et
a1．，1994；Chen，2001a，b；Wang et a1．，2005，2006)．
Buckwheat species are often classified into two groups，
the big-achene and the smal—achene group，respectively．
The former includes two cultivated species(F escIg[e72turn~
and F．tataricurn)，four natural wi ld species(F．zuogon—
gense Q．F．Chen，F．rnegaspartaniurn Q．F．Chen，F．pi—
lus Q．F．Chen and F．cymosurn Meissn)and one maD_-
ma de species(F giganteum Krotov)(Ye& Guo，1992：
Ohnishi & Matsuoka，1996；Chen，1999a，b)，while the
latter includes nine wild species，F_gracilipes(Hems1．)
Dammer ex Diels，F_leptopodum (Diels)Hedberg，F．
statice Gross，F．capillatum Ohnishi F callianthum
Ohnishi，F gilesii(Hems1)Hedberg， pleiorarnosurn
Ohnishi，F lineare(SarrL)Haraldsom and F urophyllum
Gross(Chen，1999a，b)．There are significant genetical
and morphological differences between these two groups
(Chen，1999a，b；Chen，2001；Chen et al，2004)．
It is generally believed that F．cyrnosum(一 F．cy—
Y／?OSUrn complex)is the ancestor of cultivated buckwheat，
as supported by buckwheat morphology(Hedberg，1946；
Campbell，1976)．However，based on the apparent varia—
tions among different populations of F．cymosum(==F
cymosurn complex)in，for example，morphology，cytolo—
gY，reproduction isolation and isozymes，Chen(1999a，b)
and Chen et a1．(2004)classified this species into three
separate biological species：the perennial diploid F．72e—
gaspartaniurn，perennial diploid F．pilus and perennial
allotetraploid F。cyrnosurn，colectively referred to as the
F cymosum complex． In addition，they suggested that
F．rnegaspartanium and F．pilus are the ancestors of
common buckwheat and tartary buckwheat，respectively．
To date，no reports have examined the amylase and
forma te dehydrogena se isozymes in the genus Fagopy一
九z ．In the present study，we exami ned the presence of
these two isozym es in all buckwheat species of the big-a～
chene group and some of the small-achene group at dig
ferent stages of development，in order to provide clues
and new data regarding the interspecifie phylogeny of
Fagopyrurn as well as the origin and evolution of culti—
vated buckwheat，and to provide important information
for buckwheat genetics and breeding．
1 Materials and Methods
Forty-two buckwheat accessions belonging to eight
buckwheat species were used in this study(Tables l and
2)． l accessions were provided by the Institute of
Plant Genetics and Breeding where all studies were car—
tied out． ’
1．1 Preparation of amylase(AMY)and formate dehydro-
genase(FDH。E．C 1．2．1．2)isozyme samples
The preparation of isozyme samples from sprouting
seeds largely followed the methods of Chen(1999a)and
Hu(1985)．Briefly，buckwheat seeds were placed on hu—
mid filter paper in a dish and cultured at 25℃ in an illu—
mination incubator．Ⅵ en the roots were about 2．0 C1TI
long，0．5 g of sprouting seeds were ground in extracting
solution[0．9 g poly nylpolypyrrolidone(PVPP)，0．5 g
silicon dioxide，and 0．8 mL 0．1 mol／L Tris—HC1 buffer。
pH 7．53 into uniform juice in a mortar on ice then cen—
trifuged at 9 167 g at 4℃ for 20 mi n．The supernatant
solution was used as the isozym e sample for electropho—
resis．
The preparation of isozyme samples from dry seeds
largely folowed the method of Chen(2004)．The ma in
procedures were as folows：0．3 g dry buckwheat seed
powderwere placed in a mortar with 0．9 mL 0。】mol／L
rris—HC1 bufer(pH7．5)，0．5 g silicon dioxide，and 0．96
3期 张以忠等：荞麦属植物淀粉酶和甲酸脱氢酶同功酶研究 397
g PⅥ)P，ground into uniform j uice on ice then centri—
fuged at 10 000 rpm at 4℃ for 20 min．The supernatant
solution was used as the isozyme sample for electropho—
rests．
1．2 Electrophoresis and dyeing
Polyacrylamide gel electrophoresis(PAGE)with
vertical plates was used for analysis of the amylase and
formate dehydrogenase isozymes(Chen 1999a；Chen et
a1．，2004)． The main parameters were T = 8．5
(DH8．9)and C=0。3．The eleetrophoresis buffer used in
this study was Tris—Gly(pH8．3)．The amylase and for—
mate dehydrogenase isozymes were dyed according tO Hu
398 广 西 植 物 3O卷
& Wan(1985)．
Dyeing of amylase isozyme：the gel was placed in
100 mL acetic acid buffer solution(O．15 mol／L，pH：
5．0)1．5 h at 37℃ then dyed in lOO mL dyeing solution
(0．005 g iodine and 0．079 g potassium iodide)until
white bands were showed visible． 码 e gel was then
washed with double distilled water and pictures were
taken using a data camera．
Dyeing of formate dehydrogenase isozyme：the gel
was placed in 100 mL dyeing solution(0．42 g Tris
(hydroxymethy1)aminomethane，0．14 g Sodium formate，
O．O3 g NAD Trihydrate，0．03 g NBT，and 0．06 g PMS)
for l h under dark conditions at 25℃．The gel was then
washed with double distilled water and pictures were
taken using a data camera．
1．3 Data analysis
Hierarchical cluster analysis and SPSS 1 1．1 soft—
ware were used for clustering the buckwheat accessions．
Each sample was regarded as an accession．For a certain
band，having the band noted one and otherwise zero．
The Euclidean distance，d(X ，X )，was used to estimate
the distance between any two accessions(X and XJ)：
d(X，Xi) (z ～啄 )2， ，／=1，2，3，⋯，n；志
一 l，2，3，⋯ ，m
where n number of accessions and m
of isozyme bands．
2 Results and Analysis
赢
Amy
24
22
20
18
1 6
B
14
12
1 0
8
6
4
2
1 2 3 4 5 6 7 8 9 10 1 1 12 13 1415 16 I7
十
一 自；鼻 《磷 # ％磷
《；墓 《：搿 黯 # 精
■_ 《∞
■■■ 《#， #∞ #
《搿 ●■●
自} ， 0龋 《0 ■-■ 搿 *
■_
g
r E ⋯ 目
． #∞ 《
●■■
●● —■●_ ●I ■_ ■●■■■I
●：尊 t：琦 t：船 《=嚣 《 t： c：喜 ：稿 2矬 ■_-
《黼
■_ ●I
蒲
# ■一 ■●●
- ●- ■-
● ■ ■_ ●■■
- ● ■_
●_ ●_
u #霸 一 #端 ⋯ ⋯ 自⋯ 一 《 鬻 《鲁
1 2 3 4 5 6 7 8 9 1011 121314151617
Fig．1 Amylase isozyme zymographs(A)and ideograms
(B)of sprouting seeds of buckwheat
1-17 is Es5。Es11．ES8，ES2，ES12，ZU1，PI1，GI1，lv／E4，GR1．HO1，
GIg，CY1，TA7。TAll，TA5，and TA9，respectivdy．I)ark rectangle=
very dark band，grey rectangle= dark band，empty rectangle： light
band
A total of 10 bands(AM Y-1，2，4，5，8，10，17，
= number 18，22，23)were observed
2．1 Amylase isozyme
There are no bands of amylase isozymes in al dry
seed samples but many bands in all sprouting seed sam-
ples，indicating that the amylase isozymes were not active
in dry seeds f0r all accessions，but very active in sprou—
ring seeds．Zymograms and idiograrns of sprouting seeds
from 25 buckwheat accessions belonging to eight buck—
wheat species are shown in Figs．1— 4． According to
these figures，a total of 23 amylase isozyme bands were
discovered in this study．The variations in the amylase
isozyme zymograms were significant among different
buckwheat species but were less so within the same spe—
r1 S．
esculentum accessions，
in sprouting seeds of F．
with consensus bands
AMY一1，1O，18，22，23 and varible bands Amy-2，4，
5，8，1 7 among accessions． F．esculentum var．ho—
motropicum(HO1)and F．zuogongense(ZU1)had
five bands(AMY一1，2，lO，18，23)and four bands
(AMY一1，6，l0，18)，respectively，similar to corn—
mon buckwheat．F．rnegaspartanium also had five
bands(AMY_1，4，1O，18，21)，also similar to common
buckwheat．A tota1 of eight bands(AM 1，3，4，7，10，
l1，14，g0)were observed in the F tataricum accessions，
0f which there were four consensus bands(AMIY一1，11，
14，20)and four varible bands(Amy-3，4，7，10)among ac—
cessions．F pilus(PI1)had AM_Y_l，5，1O，14，20 bands，
similar to F tataricum．F．giganteum(GI1，GI2)had a to—
tal offive bands(AM： 1，6，ll，13，19)，different from F．
cymosum(CY1)which had six bands(AMY_1，8，ll，14，
19，21)．According to these results，all big-achene group
3期 张以忠等 ：荞麦属植物淀粉酶和甲酸脱氢酶同功酶研究 399
{十
^
{哼
l ： 3 t S 7 l 哮 l岛 ll
=：l ￡ 鞠 《=盥 嚣篇 c 船
e 黜 塞 {=嚣 e= ￡= c c 潦 蓝=
《 端 ■一 e=警 《 蓝 饕一 精
《= #=警 C=嚣 =墨 端：：l C蛊尊 《=端 ￡=
■一 ■■一 ■一 赫■
■■ ■- ●_
■_ ■■- ●_
●■l
￡= =蕊 e=∞ =茹 蕊 《=嚣 ￡=璐 篇 器
l { 3 4 S 舂 7 8 挚 l诗 {
Fig．2 Amylase isozyme zymographs(A)and ideograms
(B)of sprouting seeds of F．esculentum
】一11 is respective ES1，ES8，ESI2，ES7，ES4，ES2，ES10，ES5，
ES6，ES3 and ES9．dark rectangle= very dark band，grey rectan—
gle=dark band，empty rectangle= light band
buckwheat species had one consensus band(AM Y一1)．
gracilipes(GR1)，on the other hand，had a total of
five bands(AMY．3，9，12，15，16)，very different ftom the
big-achene group of buckwheat species．
According to the distribution of bands，the amylase
isozyme zymograms could be divided into three sections：
a slow region(AM Y一1—11)，middle regi【on(AM Y-12—18)
and fast region(AMY-19—23)．The variation bands of
cormnon buckwheat al distributed at slow and middle
regions and those of tartary buckwheat only at slow re—
gion．Further，according to the amylase isozyme zymo—
grams of the different buckwheat species，four types
could be determined：Type 1 with AM Y_lO in the slow
region and AMY-18 in the middle region(F esculentum，
F．megaspartanium，F．zuogongense，and F．esculentum
var．homotropicum)；Type I1 with AMY-14 in the mi ddle
region and AMY-20 in the fast region(F．tataricum and
F pilus)；Type II1 with AⅣ【、 一11 in the slow region and
AM[Y一19 in the fast region(F．giganteum and F cymo—
SUly1)；and Wype IV with AMY-12，15，16 in the mi ddle
f+)
A
}
t 2 4 5 喜 7 孽 争 l● |l l2
谒鹾酥 辅辞释孽C #奢 ■一 譬= ￡：=l黼 嚣盘 霸
茬盘 谁鹅鹭 翻 麟 耩 鼎 姆 蝴 黼 霸 褥薛赫
镶酾薛 嚣==， 镯嘲 盘 瓣 ∞
一 ■群
■—■—■●
囊_ _ 一 I ■_ ■- ■-
I一
蠲鞫謇 奄奄麟 蝴 镬麟 瑙鳢璐 糍 ∞ ∞ =3嚣
{ ： j 5 ‘ 7 # 争 l缸 ll 豫
Fig．3 Amylase is()zyme zymographs(A)and ideograms
(B)of sprouting seeds of F．tataricum
1-12 is respective TA9，TA4，TA10，TA15，TA5，TA8，TA6，
TA7，TA12，TA2，TA14 and TA1． dark rectangle= very dark
band，grey rectangle= dark band，empty rectangle=light band
Labe
G I1
G f 2
CY1
ES5
H01
ES8
ES2
ES1
ES1
ZU1
ME4
TA5
TA9
TA1
TA7
P l 1
GR1
O 5 ’0 1 5 20 25
T T
j
． ．
．．
Fig．4 Clustering tree of the buckwheat accessions based
on amylase isozyme zymograms of sprouting seeds
region(F．gracilipes(GR1)only)
。
r ～～～～～
∞ 器¨ “ 0
—。 —。 — 。 。L— _l 。 。 【_ l_。_i E 舱 博” ∞
400 广 西 植 物 3O卷
The above buckwheat accessions were systematical—
ly clustered by means of Euclidean distance analysis(Fig．
4)．The results showed that when T was equal to 15，
the buckwheat species accessions could be divided into
five groups：Group I and Group II included the tetraploid
F．giganteum and F．cymosum，respectively~Group III，
F．esculentum，F．rnegaspartanium，F．zuogongense and
F．esculentum var．hornotropicum；Group IV，F．tataricum
and F．pilus~and Group V，F gracilipes．When T was
equal to 2O，on the other hand，the buckwheat species ac—
cessions were grouped into two groups，that is，the big-a—
chene group species and the sma ll—achene group species，
respectively，indicating that there is a distant relationship
between these two groups．
CASE
Labe
Num
0 5 1 0
ES6
ES1 2
ES3
ES7
ES9
ES1
ES1 0
ES1 1
ES2
ES4
ES8
ES5
TA1 1
TA1 3
TA1
TA8
TA1 2
TA7
TA6
TA4
TA2
TA9
TA1 0
TA1 4
TA5
20 25
Fig．5 Clustering tree of common buckwheat and
tartary buckwheat accessions based on amylase
isozyme zymograms of sprouting seeds
According to the Fig．1，the isozyme band number
of common buckwheat and tartary buckwheat in Fig。2
and 3 are easy to be defined．The 25 accessions of com—
mol buckwheat and tartary
cally clustered by means of
buckwheat were systema ti—
Euclidean distance and the
nearest neighbor analysis of hierarchical classification
(Fig．5)．It is clear that classifications for both of com—
mon buckwheat and tartary buckwheat have not obvious
relationship with their geographic distribution．The ac—
cessions native to Czech Republic(ES12)and Germany
(ESl1)are close to those native to China ，respectively．
2．2 Formate dehydrogenase isozyme
The formate dehydrogenase isozyme zymograms
and idiograms of dry seeds and sprouting seeds from 25
accessions belonging to eight buckwheat species are
shown in Figs．6 and 7．According to these figures，all
accessions had only one band，and no variations within
species or among species were observed．
3 DiSCUSSiOFI
3．1 Amylase isozyme in buckwheat and identifcation of
buckwheat species
No previous report has documented the buckwheat
amylase isozyme until now．The amylase isozyme of all
buckwheat species of the big-achene group and some
buckwheat species of the small—achene group was there—·
fore studied for the first time in the present paper．The
results showed that dry seeds have no amylase activity
while sprouting seeds were very active．The sprouting
seeds of F esculentum accessions had 10 bands including
5 consensus bands and 5 variable bands while F．tatari—
Gum accessions had 8 bands including 4 consensus bands
and 4 variable bands，less than common buckwheat．The
variations in the amylase isozyme zymograms were also
very large among different buckwheat species but were
less so within the same species．All cultivated and wild
buckwheat species used in this study had their own par—
ticular amylase isozyme zymogram，which can thus be
used for identification of buckwheat s0ecies．There are
about 16 species of buckwheat(Chen，1999a，b)，which
can be grouped two sections，that is，the big-acheen
group consisting of common buckwheat，tartary buck—
wheat，F．zuogonense，F．rnegaspartanium，F．pilus，F．
cymosum，and F giganteum； the sma ll—achene-group
consisting of 9 species(F．gracilipes，F．1eptopodum，F．
urophyllum，etc．)．The F cymosum complex consists of
three similar species that cannot be easily distinguished．
It ma y result in puzzled conclusions that the F．cymosum
complex are confused as a species． Chen(1 999a)and
Chen et a1(2004)reposed two effective methods of buck～
wheat species identification by estaerase isozyme and
3期 张以忠等 ：荞麦属植物淀粉酶和甲酸脱氢酶同功酶研究 401
glutamate oxaloacetate transaminase isozyme The results
in this study provide orie more good tool for their identi—
fication．Al species of the big-achene group had a con一
矗
sensus amylase band(AM Y-1)different from that of the
sma ll achene group，which again CaD-be used for distin—
guishing these two groups．
1 盏 3 毒 粤 螽 鑫 孽 l辔 耋重 i1 昭 蠹I 15 l舔 ” 15 19 艚 l
+ |
基江m n m ~ m= 一 _ m i ra I m m = I N
* 1 亲 露 耋 《 羹 擎 le II 12 摹 14 S 16 l窄 l暮 警 拍 II
Fig．6 Formate dehydrogenase isozyme zymographs(A)and ideograms(B)of dry seeds of buckwheat
卜21 are ES7，ES10，ES11，ES12，ZU1，PI1，GI1，GI2，ME1，ME2。ME3，ME4，ME5，ME6，ME7，GRI，HO1，CY1，TA 3·TA1
and TA15，respective．Dark rectangle=very dark band，grey rectangle—dark band，empty rectangle light band
(+)
矗
薹 3 萋 蓐 擘 警 罅 ll 霪 摹 l|垂 l蓦 描 l 糟 l謦
中 l
尊0 l==l _ —一—-_ —_ _I ：=l 黼—一—一—一—一-__一—一_!．|
l l 摹 毒 妻 蕃 亨 鼙 持 秘 琏 格 “ l薹 l‘ 籽 1． 1’
Fig．7 Formate dehydrogenase isozyme zymographs(A)and ideograms(B)of sprouting seeds of buckwheat
1．19 is ES7，ES11，ES12，ZU1，PI1，GII，GI2，ME1．ME2，ME3，ME4，ME5，ME6，GR1，HO1，CY1，TA3，TA11 and
TA13。respective．Dark rectangle= very dark band，grey rectangle=dark band，empty rectangle= light band
3．2 Formate dehydrogenase isozyme in buckwheat
As above，rio previous report has examined the for—
mate dehydrogenase isozyme of buckwheat，and this
study is the first to report this isozyrne in dry and sprou—
ring seeds of all species of the big-achene group．The re—
sults showed that activity of the formate dehydrogenase
isozyme is high in both sprouting and dry seeds．Fur—
ther，al accessions of F esculentum， tataricum，and
wild buckwheat had only one band at the same position，
indicating that the formate dehydrogenase isozyme is
highly conserved in the genus Fagopyrum．
3．3 Origin of cultivated buckwheat
Until recently，despite large variations，all perennial
natural buckwheat types with a big achene were classi—
fled as F cymosum and considered the proposed ancestor
of cultivated buckwheat(F．esculetum and F．tataricum)．
Recently，however，Chen(1999a，b，2001a，b)and Chen et
a1．(2004)put forward the concept of the F cymoxul~t
complex，classifying this single species into three differ—
ent biological species，the diploids megaspartanium
and F．pilus and allotetraploid F．cymosurn．In addition，
the morphology， chromosome karyotype， esterase
isozyme，glutama te oxaloacetate transaminase(GOT)
isozyme，int erspecific crossability and reproduction prop—
erties(Chen，1999a，b，2001a，b；Chen et a1．，2004)al1 in—
dicated that F．megaspartanium and F．pilus are the an—
402 广 西 植 物 3O卷
cestor species of F．esculentum and F．tataricum，respec—
tively．
Yamane＆ Ohnishi(2001)reported that the phylo—
genetic tree constructed using the neighhour-joining
method based on allozyme variation clarified two distinct
groups of diploid populations of the F cymosum corn～
plex．Further，Yamane et aI．(2003)showed that F ta-
taricum is similar to one member(maybe F．pilus)of the
F cymosu~n complex based on cpDNA variability．sug—
gesting that F tataricum diverged from this species in
the Tibet Hima layan area． Colectively，these reports
strongly support Chen’s(1999a)classification of F．pilus
and F megarspartanium as separate species and the hy—
potheses that they are ancestors of cultivated buck—
wheat．In this study．F．esculentum and F．tataricum had
very similar amylase isozyme zymographs to F megas—
partanium and F．pilus，respectively，further supporting
the above origin hypothesis suggested by Chen(1999a，
b)．
The results in this study showed that accession
classifications for both of common buckwheat and tartary
buckwheat have not ob~ous relationship with their geo—
graphic distribution and that the accessions native to
Czech Republic and Germany are close to those native to
China，respectively，which agree to the hypothesis that
common buckwheat and tartary buckwheat originated in
southwestern China and then spread to oversea(Chen，
199％ ，b；Ohnishi，2007)．
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