全 文 ：中国西南地区鹿药属 4 种 15 居群核型研究
?
唐自慧1 , 2 , 高云东1
??
, 周颂东1 , 何兴金1
???
(1 四川大学生命科学学院 , 四川 成都 610064; 2 成都市工业学校 , 四川 成都 610081)
摘要 : 对产于中国西南部的鹿药属 ( Maianthemum) 4 种植物进行了细胞学研究 , 包括染色体数目 , 多倍
化 , 非整倍性和随体染色体 , 以及核型不对称性和核型进化。结果表明 : 1 ) 除了在云南丽江采集的 Ma-
ianthemumtatsienensis染色体数目为 2 n= 72 之外 , 其余的居群全为 2 n = 36 ; 2 ) 核型在居群间存在变异 , 特
别是在具中部染色体和近中部染色体的数目以及随体染色体的数目和位置上。此外 , M. nanchuanense和
M . szechuanicum的核型是首次报道 , B 染色体也是首次在该属中发现。我们推测鹿药属的进化方式包括频
繁的染色体畸变以及不同水平上的多倍化 , 而中国西南部是该属的分化中心。
关键词 : 染色体 ; 进化 ; 核型 ; 鹿药属
中图分类号 : Q 942 文献标识码 : A 文章编号 : 0253 - 2700 (2009) 01 - 001 - 07
Karyotypes of Fifteen Populations of Four Species in Maianthemum
(Liliaceae) from Southwestern China
TANG Zi-Hui
1 , 2
, GAO Yun-Dong
1 * *
, ZHOU Song-Dong
1
, HE Xing-Jin
1 * **
(1 Collegeof Life Science, Sichuan University, Chengdu 610064 , China; 2 Chengdu Industry College, Chengdu 610081 , China)
Abstract: Fifteen populations of four species of Maianthemum from southwestern China were analyzed in terms of base
number, polyploidy, dysploidy, chromosome size, chromosomemorphology, satellitedchromosomes, structural heteromor-
phism, and karyotype asymmetry and karyotypeevolution . The results show that 1) chromosomal number of all the species
examined is 2 n= 36 , with M. tatsienensis (Population Lijiang, Yunnan) as an exceptional case of which the chromosomal
number is 2 n= 72 ; 2) karyotypes show difference amongpopulations, especially in the numbers of metacentric, submeta-
centric, and acrocentric chromosomes aswell as satellites . The karyotypes of M. nanchuanense and M . szechuanicum are
documented the first time . B-chromosome is discovered in this genus the first time, too . It is inferredthat the evolutionary
mechanisms in the genus Maianthemuminclude the frequent somatic chromosome aberration and the polyploidy at various
levels, and that the southwestern China is one of the differentiation centers of this genus .
Key words: Chromosome; Evolution; Karyotype; Maianthemum
Maianthemum Wiggers sensu lato ( including
Smilacina) , a genus of the tribe Polygonatae of Rus-
caceae ( Rudall et al. , 2000; APGII 2003 ) , comprises
about 35 species distributed in East Asia and North
America, with only one species M. bifolium extending
to Europe (Chen and Kawano, 2000) . This genus used
to be represented by 19 species, which occurred mainly
in the Hengduan Mountains, southwestern China ( Li ,
1990) . Karyologists havepaidmuch attention to thisge-
nus and a lot of cytological researchhas been done (Ku-
云 南 植 物 研 究 2009 , 31 (1) : 1～7
Acta Botanica Yunnanica DOI : 10 .3724?SP. J . 1143 .2009.08084
?
??
??? ?Author for correspondence; E-mail : xingjinhe@ yahoo. com. cn
Received date: 2008 - 05 - 06 , Accepted date: 2008 - 09 - 03
作者简介 : 高云东 (1985 - ) 男 , 在读研究生 , 主要从事染色体进化研究。 ?
Gao Yun-Dong and Tang Zi-Hui are equally contributed to this paper ?
Foun ?dation items: The National Natural Science Foundation of China ( 30670146 ) and the National Science and Technology Resources Project
(2005DKA21403 )
mar, 1959; Kurosawa, 1966; Sharma, 1970; Mehra
and Sachdeva, 1976; Hong and Zhu, 1987 , 1990;
Wang et al. , 1990; Wang et al. , 1993; Gu and Sun,
1998; Meng et al. , 2005 ) . 18 of the 35 species have
been studied with respect to chromosome features and
interspecific variation in karyotype of somatic chromo-
somes (Meng et al. , 2005) .Theorigin and evolutionof
Maianthemumhavebeendiscussedwith evidences from
cytological studies ( Hong and Zhu, 1987 , 1990) .
This paper provides results of our study on karyo-
type analysesof somatic chromosomes in 15 populations
of four speciesof Maianthemumfromsouthwestern Chi-
na, including 2 species not studied before, and dis-
cusses the karyotype evolution according to both our
and previous research data .
1 Materials and methods
The species names and populations, along with their ori-
gins, are list inTable1 . Voucher specimens aredeposited in the
herbarium of Sichuan University (SZ) . Plant individuals were
collected fromnativehabitats, and cultivated in pots for cytologi-
cal study . Root tipswerecollected in themorning from each in-
dividual , pretreated with a solutionof mixed liquor (0 .1 % aque-
ous colchicines: saturated dichlorobenzene= 1∶1 ) for 6 - 7hr at
20 - 21℃ , fixed in 3∶1 ethanol-acetic acid for at least one day,
and stored in70% ethanol at 4±2℃ . The root tipsweremacer-
ated in amixture of 1 mol?L HCl for 10 min at 60℃ , stained by
1 % acetic orcein for 2 - 3 hr, and squashed on a glass slide to
observe thecells . Onetofive individual plants fromeachpopula-
tion and several roots per plant were studied . The chromosomes
of at least 30 metaphase plates from each root were studied .
Number, size and shape of chromosomes were observed, and
karyotype asymmetrywas evaluated . Satelliteswere not taken into
calculation of chromosomesizeandthe highest number of satellit-
ed chromosomes in each plate was recorded, followed Lifante
(1996) . Nomenclatures for the centromeric positions of chromo-
some introduced by Levan, Fredga and Sandberg ( 1964 ) were
followed: m= median centromeric chromosome with arm ratio of
1 .0 - 1 .7 ; sm= submedian centromeric chromosomewitharmra-
tio of 1 .7 - 3 .0 ; st = subterminal centromeric chromosome with
armratio of 3 .0 - 7 .0 . Karyotype asymmetry was described ac-
cording to Stebbins (1971 ) , who establishedthese by recognizing
three degrees of difference (A-C) between the largest and small-
est Chromosome of the complement, and four degrees ( 1 - 4 )
with respect to the proportion of chromosomeswhicharemetacen-
tricwith an arm ratio of less than 2∶1 . So there are 12 types .
The index of asymmetry (As K% ) also adopted Arano (1963 ) ,
i . e . the ratio of the sum of long arm length to the total haploid
length of the chromosome complement .
2 Results and Discussion
The karyotype structures, classifications and
asymmetry indexesof all populationsof thespecies ana-
lyzed are list in Table 2 . Diploid was found in all taxa
studied, while tetraploid found only in Maianthemum
tatsienensis, and dysploidy ( 2 n = 35) was found in a
Yunnan population of M. henryi . Most taxa have a
karyotype asymmetry of type 2C according to Stebbins
(1971) , except M. tatsienensishas a2B type .The in-
dexes of asymmetry ( As K % ) of these species are
high, which ranges from 64 .75% - 66 .82% . The bi-
modal karyotype was obvious .
Table 1 Origins, altitude and voucher numbers of the Maianthemumspecies and populations examined
Taxon Locality Altitude ( m) Collector & Voucher
Emei J inding, Sichuan 3070 ?Z .H . Tang, SZTM-017
Emei , Leidongping, Sichuan 2430 ?Z .H . Tang, SZTM-158
Emei , Chudian, Sichuan 1740 ?Z .H . Tang, SZTM-206
M ?. tatsienensis ( Franchet) LaFrankie J infoshanzhuang, Chongqing 1800 Z .H . Tang, SZ TJ -06
Guofodong, J infoShan, Chongqing 2000 ?Z .H . Tang, SZ TJ -11
Wanyuan, Dazhou, Sichuan 1850 ?Z .H . Tang, SZ TW-01
Lijiang, Yunnan 2800 ?Q . Wang, SZ WQ-006 *
Dali , Yunnan 2930 ?Q . Wang, SZ WQ-003 *
M ?. henryi ( Baker) LaFrankie Emei Chudian, Sichuan 1740 Z .H . Tang, SZ TM-11
Kangding, Sichuan 2880 ?Q . Wang, SZ WQ-0027 O
Wolong, Sichuan 3400 ?Q . Wang, SZ WQ-0026 O
M ?. szechuanicum ( F <. T . Wang & Tang) H . Li Emei Leidongping, Sichuan 2430 Z .H . Tang, SZ TM-008
M ?. atropurpureum ( Franchet) LaFrankie Emei Luohanpo, Sichuan 2300 Z .H . Tang, SZ TM-040
Guofodong, JinfoShan, Chongqing 2000 ?Z .H . Tang, SZ TJ -01
M ?. nanchuanense H . Li & J . L . Huang J infoshanzhuang, Chongqing 1800 Z .H . Tang, SZ TJ -05
2 云 南 植 物 研 究 31 卷
Table 2 Karytotype structures of populations and species of Maianthemum
Taxon Collection number Karyotype formula As K % Type
M ?. tatsienensis ( Franchet) LaFrankie SZ TM-017 2 n= 36 = 14m+ 14sm+ 8st 65 .80 2 PB
SZ TM-158 ?2 n= 36 = 18m(1sat) + 12sm( 2sat) + 6st 66 .05 2 PB
SZ TM-206 ?2 n= 36 = 20m+ 12sm+ 4st 65 .00 2 PB
SZ TJ-06 D2 ?n= 36 = 14m+ 18sm+ 4st 64 .73 2 PB
SZ TJ-11 D2 ?n= 36 = 12m+ 18sm+ 6st 65 .85 2 PB
SZ TW-01 j2 ?n= 36 = 12m+ 20sm+ 4st ( 1sat) 65 .39 2 PB
SZ WQ-006 ?2 n= 72 = 32m(1sat) + 28sm+ 12st 64 .75 2 PB
M ?. henryi ( Baker) LaFrankie SZ WQ-003 2 n= 36 = 14m+ 14sm( 1sat) + 8st 66 .15 2 PC
SZ TM-11 b2 ?n= 36 = 18m+ 12sm+ 6st 66 .04 2 PC
SZ WQ-0027 ?2 n= 36 = 18m+ 10sm+ 8st 66 .28 2 PC
SZ WQ-0026 ?2 n= 36 = 20m+ 6sm+ 10st 66 .48 2 PC
M ?. szechuanicum ( F <. T . Wang & Tang) H . Li SZ TM-008 2 n= 36 = 14m+ 14sm+ 8st 66 .80 2 PC
M ?. atropurpureum ( Franchet) LaFrankie SZ TM-040 2 n= 36 = 20m+ 8sm(1sat) + 8st 65 .37 2 PC
SZ TJ-01 D2 ?n= 36 = 16m+ 16sm+ 4st 65 .59 2 PC
M ?. nanchuanense H . Li & J . L . Huang SZ TJ-05 D2 n= 36 = 22m(2sat) + 2sm+ 12st + B 66 .28 2 PC
2 .1 Base number
The chromosome number of 2 n= 35, 36 , 72 has
been found in our study . Kumar ( 1959 ) , Kurosawa
(1966) , and Sharma (1970 ) reported the chromosome
number 2 n = 28, 36 , 66 , 72 . Mehra and Sachdeva
(1976 ) reported the chromosome number 2 n = 36 of
S. purpurea . The chromosome number of S. japonica
and S. henryi collected in Taibai mountain of China is
2 n= 36 . Hong and Zhu (1987) , Wang et al . (1990)
reported the chromosome number 2 n = 36 of
S. tatsienensis collected from Sichuan province . Wang
et al . (1993) reported the chromosome number 2 n= 36
of somespeciesof Smilacina fromYunnan province . Gu
and Sun (1998) reported the chromosome number 2 n=
54 of S. fusa fromMotuo Xizang (Tibet) . It could be
inferred that most species of Maianthemumhas a chro-
mosomal number of 2 n = 36 , occasionally 2 n = 54 or
72 . All the studied species in Maianthemumhavechro-
mosome number of 2 n = 36 , which is currently consid-
ered as diploid . Our research confirmed thishypothesis,
becausewedidn′t find any individual has a chromosome
number below 36 (except a dysploidy with 2 n = 35 ) .
The base number 2 n = 18 was very high, and it was
against with Stebbins′(1971) hypothesis that all genera
and families should have a base number equal or below
x= 12 . So this group might be an ancient polyploidy .
Further studies need to confirmthis hypothesis .
2 . 2 Polyploidy and dysploidy
Hong and Zhu ( 1987 ) have shown an incidence
of polyploidy in Maianthemum, and this is confirmed
in our study . By putting thesedata together with infor-
mation reported in earlier studies, the diploid level has
been found in all taxa studied, and the tetraploid level
found in M. tatsienensis and M. atropurpureum . M. f-
uscumwith 2 n= 54 may be an ancient hexaploid with
base number x= 9 , as discussed above .
The number 2 n= 35 was only found in a Yunnan
population of M. henryi , which makes up 6% of the
total cells examined (Figs. 9 - 10) . The largest chro-
mosome is ametacentric of typeM , with double centro-
meres, which could have resulted from a Robertsonian
translocation between twosmchromosomes and the loss
of the short arms (Yu and Huang, 1994 ) . The ratio of
those cells with 2 n = 35 was low, further studies are
needed to determine its role .
2 . 3 Polymorphism of karyotype
There are significant inter-population differences
in karyotype, particularly in B chromosomes, as well
as the number and location of satellites and karyotype
asymmetry . Moreover, our karyotypedata areoftendif-
ferent fromthose reported in the earlier studies .
2 . 3 .1 B chromosome B chromosome is found in
Maianthemumthe first time, andonly present in M. n-
anchuanense, and B chromosome was observed only in
one individual among the three individuals studied by
us . According to the ninth pair chromosome ( Figs. 25
- 26) , wethink thepossiblemechanismis chromosomal
structure heteromorphism in response to environmental
31 期 TANG Zi-Hui et al. : Karyotypes of Fifteen Populations of four Species in Maianthemum (Liliaceae) . . .
stresses . It seems theB chromosome has somerelativity
withmorphogenesis, such as the size of rhizome, stem
and inflorescence; whether those are just individual
variation or intrinsic relation need further experiment to
work it out, as well as the influences of the fertility .
2 . 3 .2 Bimodality The karyotypes of 15 populations
studied are all bimodal . Ten pairs of chromosomes in
the karyotype are significantly larger than the rest, and
the number of themacrochromosomes is almost equal to
that of microchromosome . We agree with Levitzky
(1931) that bimodal karyotypes comefromunequal trans-
location . Besides, centric fusions could have played an
important roleinthe evolutionof thekaryotypeof thisge-
nus . Bimodality is considered as an extremely special-
ized form of asymmetry ( Stebbins, 1971 ) , so the ge-
nus Maianthemum is likely to be an advanced group in
the tribe Polygonatae of Ruscaceae .
2 . 3 .3 Chromosomal structural heteromorphism
There is a high level of structural heteromorphism
present in both satellited and non-satellited chromosomes
Figs . 1 - 8 . Photomicrographs of metaphase chromosomes and their karyotypes in four populations of M. tatsienensis . 1 - 2 . Popula-
tionEmei , Leidongping, Sichuan, 2 n= 36; 3 - 4 . Population Gufodong, Chongqing, 2 n= 36; 5 - 6 . PopulationWang yuan Sichuan,
2 n= 36; 7 - 8 . Population Li jiang Yunnan, 2 n= 72 . ( ( Arrows indicating satellited chromosome . Scalebars= 100μm)
4 云 南 植 物 研 究 31 卷
Figs . 9 - 18 . Photomicrographs of metaphase chromosomes and their karyotypes in four populations of M. henryi . 9 - 10 . Population Dali , Yun-
nan, 2 n= 35; 11 - 12 . Population Dali , Yunnan 2 n= 36; 13 - 14 . Population Emei , Chudian, Sichuan, 2 n= 36; 15 - 16 . Population Kang-
ding, Sichuan, 2 n= 36; 17 - 18 . Population Wolong, Sichuan, 2 n= 36 . ( Arrows indicating satellited chromosome . Scalebars= 100μm)
in almost all species . Chromosomal structural hetero-
morphism occurs frequently in M. henryi with double
centromeres, while M. tatsienensis has the lowest lev-
el . Chromosomal heteromorphismis obvious, which in-
dicates that a karyotypic differentiation does exist
among the populations . We havereason to believe that
pericentric inversion and the Robertsonian or unequal
reciprocal translocations also exist . Hence the karyo-
type is somewhat polymorphic at the intraspecific level .
We believe that the diversified habitats might have in-
duced the chromosomal structural heteromorphism, and
that the facultative apomictic reproduction and
51 期 TANG Zi-Hui et al. : Karyotypes of Fifteen Populations of four Species in Maianthemum (Liliaceae) . . .
polyploidy might help to surmount the obstacle of re-
duced fertility and thus maintain chromosomal hetero-
morphismamong populations (Yang et al. , 1992) .
3 Differentiation center
Firstly, in all the 19 species of Maianthemum in
China, eight are concentrated in HengduanMountains,
withothers distribute sparsely in the rest of East Asia
and mostly have a distribution in southwestern China .
This distribution pattern shows that Hengduan Moun-
tains, located in the centre of southwestern China,
have a core status in thedistributionof Maianthemum .
Figs . 19 - 26 . Photomicrographs of metaphase chromosomesand their karyotypes in three species of Maianthemu . 19 - 20 . M . szechuanicum, 2 n
= 36 ; 21 - 24 . M . atropurpureum, 2 n = 36 . 21 - 22 . Population Luohanpo, Sichuan, 2 n = 36; 23 - 24 . Population J infoshanzhuang,
Chongqing, 2 n= 36 ; 25 - 26 . M. nanchuanense . ( Arrows indicating satellited chromosome . Scale bars= 100μm)
6 云 南 植 物 研 究 31 卷
Secondly, as discussed above, the karyotype evolution
of Maianthemum includes polyploidy, structural reor-
ganizations as well as dysploidy . Structural reorganiza-
tions canproduce new supergene, sothat it will begiv-
en a higher fitness to occupy a new niche, while the
structural reorganizations can be fixed through asexual
reproduction . Thosevariations in present work indicate
that thisgenus areyoung and areundergoingan intense
differentiation progress .
On the other hand, according to previous studies
( Kumar, 1959; Kurosawa, 1966; Sharma, 1970;
Mehra and Sachdeva, 1976; Gu and Sun, 1998) and
the present work, it is noteworthy that all the polyploid
taxa are concentrated in the Hengduan Mountains re-
gion, which is recognized as a biodiversity‘hot spot’
(Myers et al. , 2000 ) , and hold the richest temperate
flora of seed plants in the world ( Wu, 1988; Li and
Li , 1993) . Floristically, it is also a highly natural re-
gion and is one of the areas of the world with a high
concentration of endemic species ( Wu, 1988; Li and
Li , 1993) . So the origin and the evolvement of ploidy
level are probably due to the geological events which
created different habitats .
In a word, the correlation of modern distribution
pattern and the polymorphism of karyotype observed in
present study, together with theparticularity of Hengd-
uan Mountains discussed above show that southwestern
China is probably the diversity and differentiation cen-
ter of genus Maianthemum .
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