Journal of Genetics and Genomics (Formerly Acta Genetica Sinica) February 2007, 34(2): 171-179
Research Article
Genetic Diversity of Volatile Components in Xinjiang Wild Apple (Malus sieversii) Xuesen Chen①, Tao Feng, Yanmin Zhang, Tianming He, Jianrong Feng, Chunyu Zhang Biological Laboratory of Pomology, Shandong Agricultural University, Tai’an 271018, China
Abstract: To evaluate genetic relationships using qualitative and/or quantitative differentiation of volatile components in Xinjiang Wild Apple (Malus sieversii (Lebed.) Roem.) and to acquire basic data for the conservation and utilization of the species, aroma components in ripe fruit of M. sieversii obtained from 30 seedlings at Mohe, Gongliu County, Xinjiang Autonomic Region, China, and in ripe fruit of 4 M. pumila cultivars (‘Ralls’, ‘Delicious’, ‘Golden Delicious’, and ‘Fuji’) were analyzed using head space-solid phase microextraction and gas chromatography-mass spectrometry. The results indicated that the values of similarity coefficient concerning volatile types between the two species were in accordance with the evolution of M. pumila cultivars (forms), and that M. sieversii seedlings showed considerable genetic variations in these aspects: the total content of volatile components, the classes and contents of each compound classes, the segregation ratio, and content of main components. The results showed significant difference among seedlings and wide genetic diversity within the populations. Comparison of the volatile components in M. sieversii with those in M. pumila cultivars showed that the common compounds whose number were larger than five with the contents over 0.04 mg/L simultaneously between M. sieversii and M. pumila cultivars belonged to esters, alcohols, aldehydes or ketones. This suggests fundamental identity in main volatile components of M. sieversii and M. pumila cultivars. The results above sustained the conclusion “M. sieversii is probably the ancestor of M. pumila”. However, there were 48 compounds present in M. pumila that were not detected in M. sieversii, including 6 character impact components (i.e., propyl acetate, (Z)-3-hexenal, 2-methyl-1-butanol acetate, pentyl acetate, 3-furanmethanol, and benzene acetaldehyde). This suggested that in the domestication of M. pumila, introgression of other apple species, except for M. sieversii, by interspecies hybridization was possible. There were 177 compounds in total belonging to 11 classes detected in 30 M. sieversii seedlings, including esters, alcohols, ketones, aldehydes, acids, benzene ramifications, terpenes, heterocycles, hydrocarbon derivates, acetals, and lactones. Among them, acetals and lactones were not detected in M. pumila cultivars, 90 compounds were unique to M. sieversii, and 7 components (1-butanol, ethyl butanoate, 1-hexanol, ethyl hexanoate, 3-octen-1-ol, ethyl octanoate, and damascenone) belonged to character impact odors. Thus, the potential of M. sieversii in “utilization conservation” is enormous as a rare germplasm on genetic improvement of M. pumila cultivars. Keywords: Malus sieversii; Malus pumila; volatile components; genetic diversity
As a member of the Rosaceae, apple (Malus
apple is also one of the competitive industries for the
pumila Mill.) ranks the most agronomically important
farmers in the wide region of northern and northwest-
fruit tree in temperate zone. Apple has been listed as
ern China. However, among the more than 1,000 ap-
one of the 11 types of predominant production by Ag-
ple cultivars (strains) created by worldwide breeders
ricultural Ministry of China, in addition cultivation of
in the last 50 years, a few right-protected apple culti-
Received: 2006-02-14; Accepted: 2006-04-04 This work was supported by National Natural Sciences Foundation of China (No. 30471196). ① Corresponding author. E-mail:
[email protected]; Tel: +86-538-824 9338; Fax: +86-538-824 2364 www.jgenetgenomics.org
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Journal of Genetics and Genomics
vars are commercially cultivated in China. China began to focus on apple breeding program to meet the cultivar need of apple production. In worldwide apple breeding, there is an interesting phenomenon that about 70% to 80% of parents of over 1,000 cultivars are such “Founder parents” as ‘Ralls’, ‘Fuji’, ‘Delicious’, and ‘Golden Delicious’. Inbreeding between founder parents inevitably leads to narrow genetic background, lower adaptability, and resistance to dis-
eases and insects of current apple cultivars[1 3]. Therefore, establishment of a novel apple breeding system, which combines modern biotechnology with traditional sexual crossing is of great significance for sustainable development of apple industry. However, distant hybridization is an efficient way to create new germplasm, which widens genetic basis to crops including apple. China is one of the oldest breeding centers of Malus plants and has extremely abundant germplasm resources of apple. There are 21 wild Malus species that originated from China, among which M. sieversii (Lebed.) Roem. is believed to be progenitor of modern cultivated apple[4]. M. sieversii is mainly distributed in Tianshan Mountains of Central Asia including the Ily Valley of West China, the State Alma-Ata and Taldy-Kurgan of Kazakhstan, and the State Issyk of Kyrgyzstan. In 1989, 1993, 1995, and 1996, an expedition of Cornell University of USA explored M. sieversii. in Kazakhstan, Kyrgyzsatan, and Uzbekistan, and collected open-pollinated materials (seeds). Consequently, a germplasm garden and gene bank of M. sieversii was established in Agricultural Research Institute, Geneva. Since 2002, distant hybridization between M. sieversii and M pumila cv ‘Gala’ has been done with an objective to create new apple cultivars with excellent quality and great resistance[5,6]. The wild fruit forest in Ily, which locates on the slopes of the Ily Valley, one of important sections of central Asia Tianshan Mountains wild fruit forest, was considered as a peculiar “Ocean climate” broadleaf tree forest type in Central Asia desert climate[7]. In the
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Ily Valley, M. sieversii forms forest in which it is the dominant tree. The total area of M. sieversii in the Ily Valley is 9,330 hm2. Although Chinese government had established a nature conservation region in the Ily Valley to protect wild species including M. sieversii, but the area of M. sieversii is reducing because of overgrazing, uncontrolled cutting, and insect damage caused by Agrilus mali Mats. It is essential to take practical and effective measures to conserve the valuable genetic germplasm. In the past decades, studies on geographical distribution, variable forms, cytological observations concerning M. sieversii have -
been done[8 13]. Till now, not much knowledge has been readily available to researchers in China about population structure, genetic diversity, and reproduction system of M. sieversii. In 2004–2005, M. sieversii in the Ily Valley was investigated and 500 germplasm accessions were collected. A systematic study and evaluation is in process, which includes morphology, chemistry (compositions of sugars and acids, mineral elements, etc.), and molecular systematics.To provide basic data for the origin and evolution of domesticated apple, the genetic variation and diversity of volatile components in M. sieversii were analyzed for the first time in the present study using head space-solid phase microextraction (HS-SPME) and gas chromatography-mass spectrometry (GC-MS). The results would provide scientific basis for conservation and utilization of M. sieversii.
1 1. 1
Materials and Methods Materials
From late August to early September 2004 and 2005, about 2 kg ripe fruits from 30 randomly selected trees were collected from the wild M. sieversii population located in Mohe (N43°15, E82°51, the altitude 1,290-1,330 m ), Gongliu County, Xinjiang Autonomous Region. As an outcrossing species, the differences among M. sieversii seedlings mostly resulted from genetic variation. Additionally, a cultiwww.jgenetgenomics.org
Xuesen Chen et al.: Genetic Diversity of Volatile Components in Xinjiang Wild Apple (Malus sieversii)
vated apple cultivar ‘Erqiuzi’ and an apple form ‘Haitangguo’ were also collected at Gongliu County. The controlled M. pumila cultivars ‘Golden Delicious’, ‘Delicious’, ‘Ralls’, and ‘Fuji’ were collected in Tai’an, Shandong Province. All the samples were transported to the laboratory for extraction and detection of volatile components. The experimental test was done at the Biological Laboratory of Pomology, Shandong Agricultural University and Key Laboratory of Industrial Biotechnology, Ministry of Education, Southern Yangtze University. 1. 2
Methods
1.2.1
Extraction of volatile components
The exposed fiber coated with polydimethylsiloxane (PDMS, 100 μm thick) was manually preconditioned at 270℃ for 1 h in the GC injection port. First, a washed fruit was peeled off and then stones were removed. Immediately its juice was extracted with a juice extractor. A total of 7 microlitters of the juice were transferred to a vessel. Then, 2.0 g NaCl and 1.0 mL 3-Octanol (0.68 mg/mL) were added to the vessel. The final concentration of 3-Octanol was 0.09714 mg/L. The sample container was then placed on a magnetic stirrer. After been pushed in the sample container, the fiber was extended and was exposed to the sorption surface above the liquid surface for sam-
34℃ for 3 min, 34℃ to 50℃ (3℃/min), 50℃ to 140℃ (6℃/min), then 140℃ to 230℃ (10℃/min), and 230℃ (4 min). The temperature of the injector was 250℃. A nondiffluent mode was used with a sample size of 1 μL. The interface between GC and MS is at 250℃. Electron impact ionization was at 70 eV and injecting current was 200 uA. A full scan mode was used. 1.2.3
Qualitative and quantitative analysis
Statistical analyses were done using the software Xcalibur. The qualitative analysis was as follow: spectrometric data were compared with those obtained from the NIST HP59943C original library mass-spectra (Hewlett-Pachard) and Wiley library, combined with manual resolution of mass spectra and relative reports. Only results identified with positive and negative matching values more than 800 (maximum is 1,000) are shown in this article. A quantitative analysis was done with 3-Octanol (Sigma Ltd. Co., Louis, USA) as internal standard (0.09714 mg/L). To estimate quantitative variation of volatile components, a statistical index, CV was used. The index is calculated from the formula given below: CV=
is the mean of a sample.
duced into the GC injector where the desorption was
1.2.4
carried out at 270℃ for 5 min. Conditions of gas chromatography-mass spectrometry
The identification and quantification of volatile components in samples were done on a Finnigan Trace GC-MS using the method described by Wang et al.[14] and Chen et al.[15] with a slight modification. Separations were done with a Supelco CV1701 column, which was preconditioned at 300℃ for 2 h. Helium was used as the carrier gas (0.8 mL/min). The analysis was conducted following the program at www.jgenetgenomics.org
s ×100 x
where s is the standard deviation of samples and x
pling (at 40℃) for 35 min. The fiber was then intro-
1.2.2
173
Units of odor and characteristic impact components
The unit of odor (Uo) is the ratio of the content of a particular compound to its threshold value. If the value of Uo is larger than 1, the volatile contributes to the whole aroma and the volatile component belongs to character impact components. 1.2.5
Relationship analysis
Similarity coefficient (Sc) is used to test the relationship between M. sieversii and domesticated apple cultivars. The index is calculated from the formula given below:
174
Sc =
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with an average of 2.0229 mg/L and a higher variation coefficient of 38.94% (Table 1).
C ×100 a+b
where C is the number of common volatile compounds between M. sieversii and domesticated apple cultivar, a is the number of volatile compounds of M. sieversii and b is the number of volatile compounds of domesticated apple cultivar.
2
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Results
2. 1
Genetic variation and genetic diversity of volatile compositions in M. sieversii
2.1.1
Genetic variation of types and total content of volatile compounds in M. sieversii seedlings
The detected information of volatile compounds in the 30 M. sieversii accessions was subjected to statistical analysis. A total of 177 volatile compounds were identified and quantified in the 30 seedlings. The numbers of volatile compounds ranged from 44 to 54
2.1.2
Variation of types and concentration of volatile compounds in M. sieversii accessions
Volatile compounds that were identified and quantified are shown in Table 1. Based on the chemical structure, the 177 volatile compounds detected in M. sieversii accessions were classified and analyzed. There were namely 56 esters, 36 alcohols, 29 ketones, 16 aldehydes, 6 acids, 11 benzene ramifications, 8 heterocycles, 4 hydrocarbon ramifications, 4 lactones, 4 terpenes, and 3 acetals. The 30 accessions of M. sieversii showed significant difference with the variation coefficients of 23.8%-164.0% in each type of chemical class as well as that of 42.9%-411.5% in content. 2.1.3 Content variation of main volatiles in M. sieversii seedlings
with an average of 49 per accession. There was a
Twenty-five main volatile compounds (with
slight variation among seedlings with a variation co-
content more than 0.1 mg/L) are listed in Table 2, which include 11 esters, 6 alcohols, 6 aldehydes, 1
efficient of 5.46%. However, these seedlings showed significant difference in total content of volatile compounds, ranging from 0.8038 to 4.1435 mg/L Table 1
benzene ramifications, and 1 acetal. Regarding the index “presence: absence” separation ratio of volatile
Comparison of categories and contents of aroma components in Malus sieversii and M. pumila M. sieversii Types
Compounds
M. pumila
Content (mg/L)
Variation Mean range
CV (%)
Esters
15.8
9-25
Alcohols
11.5
Aldehydes
Types
Mean
Variation range
CV (%)
Mean
27.2
0.5536
0.0638-1.6966
60.6
15.5
9-26
8-18
20.9
0.7007
0.2953-1.9327
59.5
9.5
5.9
4-9
24.1
0.6414
0.0825-1.2389
42.9
Ketones
7.0
4-11
24.9
0.0402
0.0112-0.0876
Terpenes
2.1
1-4
23.8
0.0187
Benzene ramifications
2.6
1-6
53.2
Heterocycles
1.3
0-4
76.0
Hydrocarbon derivates
0.6
Acids
Content (mg/L)
Variation CV (%) range
Mean
Variation range
CV (%)
48.99
0.3413
0.0634-0.6544
89.55
5-14
44.24
0.2976
0.2449-0.3797
21.76
8.8
7-11
19.52
1.4881
1.0062-2.3383
39.59
50.4
6.0
3-10
49.07
0.0418
0.0104-0.1334
145.92
0.0017-0.0726
114.6
0.8
0-1
66.67
0.0014
0-0.0020
67.35
0.0062
0.0012-0.0181
66.9
3.0
2-5
47.14
0.0256
0.0108-0.0408
64.10
0.0033
0-0.0261
146.3
1.0
0-3
141.42
0.0384
0.0045-0.1188
140.26
0-3
128.4 0.0165
0-0.3546
411.5
0.3
0-1
200.00
0.0003
0-0.0010
200.00
0.3
0-2
164.0 0.0027
0-0.0274
244.0
4.0
1-6
54.01
0.0249
0-0.0761
139.47
Acetals
0.8
0-3
89.6
0.0385
0-0.1550
121. 7
0
0
0
0
0
0
Lactones
0.5
0-2
134.8 0.0011
0-0.0124
228.1
0
0
0
0
0
0
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Xuesen Chen et al.: Genetic Diversity of Volatile Components in Xinjiang Wild Apple (Malus sieversii)
Table 2
The main volatile compounds in Malus sieversii
Categories Alcohols
Esters
Aldehydes
HR Acetals
175
Compounds
Formula
Segregation ratio (presence: absence)
Contents(mg/L) Mean
Variation range
CV (%)
Ethanol
C2H6O
30:0
0.2196
0.0391-0.3250
29.90
1-butanol
C4H9O
15:15
0.1078
0-0.5780
140.36
2-methyl-1-butanol
C5H12O
17:13
0.0165
0-0.1371
171.69
1-hexanol
C6H14O
29:1
0.1602
0-0.7202
93.78
1-octanol
C8H18O
7:23
0.0056
0-0.1515
493.85
3,4,5-trimethyl-4-heptanol
C10H22O
30:0
0.1250
0.0150-0.5246
94.42
Ethyl acetate
C4H8O2
29:1
0.0998
0-0.8160
172.22
Butanoic acid, ethyl ester
C6H12O2
30:0
0.0623
0.0016-0.1975
88.81
1-butanol, 2-methyl-, acetate
C7H14O2
3:27
0.0004
0-0.0054
327.59
Butanoic acid, 3-hydroxy-, methyl ester
C5H10O3
11:19
0.0146
0-0.4062
506.03
Butanoic acid, 3-hydroxy-, ethyl ester
C6H12O3
26:4
0.1825
0-0.4880
73.92
Ethyl hexanoate
C8H16O2
3:27
0.0192
0-0.3225
376.92
Hexyl acetate
C8H16O2
6:24
0.0022
0-0.0226
262.70
Butyl hexanoate
C10H20O2
2:28
0.0072
0-0.2094
533.18
Hexyl hexanoate
C12H24O2
10:20
0.0080
0-0.1527
360.06
Pentanoic acid, 3-hydroxy-, ethyl ester
C7H14O3
19:11
0.0414
0-0.2950
176.16
3-hydroxy-, ethyl dodecanoate
C14H28O3
3:27
0.0162
0-0.2588
377.58
5-methyl-2-furfural
C6H6O2
8:22
0.0013
0-0.0148
271.98
(E)-2-hexenal
C6H10O
30:0
0.3633
0.0569-0.6627
45.42
(E,E)-2,4-hexadienal
C6H8O
30:0
0.0317
0.0049-0.0668
49.29
Furfural
C5H4O2
1:29
0.0012
0-0.0365
547.72
5-(hydroxymethyl)-2-furaldehyde
C5H11O2
7:23
0.0034
0-0.0532
324.06
Hexanal
C6H12O
30:0
0.2287
0.0156-0.5257
52.60
cis-2,3-epoxyoctane
C8H16O2
2:28
0.0155
0-0.3445
425.40
2,4,5-trimethyl-1,3-dioxolane
C6H12O2
19:11
0.0368
0-0.1550
127.12
compounds, an obvious difference was found in these
with the variation coefficients of 29.90% to 547.72%.
seedlings. Among the 25 compounds, only 6 volatiles
2. 2
Comparison of volatile compounds between M. sieversii and M. pumila cultivars (forms)
tanoate, (E)-2-hexenal, (E,E)-2,4-hexadienal, and
2.2.1
Comparison of types and contents of volatiles
hexanal) were detected in all the 30 seedlings with a
In Table 1, the common compounds whose number were larger than five with the contents over 0.04 mg/L simultaneously between M. sieversii and M. pumila cultivars belonged to esters, alcohols, aldehydes or ketones. This suggests a basic identity between the main volatile compounds of M. sieversii
(i.e., ethanol, 3,4,5-trimethyl-4-heptanol, ethyl bu-
separation ratio of 30:0, whereas the separation ratios of other 19 compounds were 1:1, 1:2, 1:3, 1:4, 1:9, 1:14, 1:29, 2:1, and 3:1. Furthermore, extensive variation in types and contents of the main volatile compounds was also presented among the 30 seedlings www.jgenetgenomics.org
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Journal of Genetics and Genomics
and M. pumila cultivars. However, M. sieversii contained 11 classes of volatiles (i.e., esters, alcohols, ketones, aldehydes, acids, benzene ramifications, heterocycles, hydrocarbon ramifications, lactones, terpenes, and acetals). While M. pumila cultivars contained 9 classes of volatile (i.e., esters, alcohols, ketones, aldehydes, acids, benzene ramifications, heterocycles, hydrocarbon ramifications, and terpenes). Lactones and acetals were not detected in M. pumila cultivars. 2.2.2
Similarity coefficient between M. sieversii and M. pumila
In the cultivated apples that were assayed, ‘Erquizi’ is the oldest cultivar developed in the former Soviet Union in the 18th century and widely cultivated in Xinjiang Autonomous Region. ‘Haitangguo’ is an intermediate form between M. sieversii and M. pumila, which was usually a tree that is planted on the roadsides. The cultivars ‘Delicious’, ‘Golden Delicious’, and ‘Ralls’ were selected from seedlings (whose parents were unsure or unknown) in U.S. in 1887, 1888, and 1905, respectively. ‘Fuji’ was selected from 6,000 offspring resulting from the crossing of ‘Ralls’בDelicious’ in 1962. In the present study, similarity coefficient of common volatile compounds was introduced to evaluate the genetic relationship between each apple cultivar (form) and M. sieversii. As shown in Fig. 1, the greatest similarity (21.12) was found between M. sieversii and the oldest apple cultivar ‘Erquizi’, whereas the least similarity (13.51)
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was found between M. sieversii and the recently developed apple cultivar ‘Fuji’. The result suggested that the similarity coefficient was in accordance with the evolution of apple cultivars (form). 2.2.3
Volatile composition and character impact odors
A total of 177 volatiles were detected in 30 M. sieversii seedlings and 135 volatiles in 4 M. pumila cultivars, with 87 common in both species. Among them, 90 compounds were unique to M. sieversii and 48 to M. pumila. Table 3 shows the 17 character impact odors in 30 M. sieversii accessions and 4 M. pumila cultivars. The four volatiles including hexanal, 2-methy-ethyl butanoate, (E)-2-hexenal, and hexyl acetate were common to the two apple species with the unit of odor ranging from 1.26 to 81.13. There were seven compounds, such as 1-butanol, ethyl butanoate, 1-hexanol, ethyl hexanoate, 3-octen1-ol, ethyl octanoate, and damascenone, unique to 30 M. sieversii accessions. A significant difference was found in mean odor units, ranges of odor units and, ratios of seedlings, which possess character impact odors. Six compounds (i.e., propyl acetate, (Z)-3hexenal, 2-methyl-1-butanol acetate, pentyl acetate, 3-furanmethanol, and benzene acetaldehyde) possessed unique character impact odors of M. pumila cultivars, and the odor unit ranged from 1.27 to 177.2.
3
Discussion
3. 1
Genetic diversity of volatile compounds in M. sieversii and origin and evolution of M. pumila
Ily wild fruit forest, which mainly consists of M. sieversii, Armeniaca vulgaris, and Juglans regia, was considered as a peculiar “Ocean climate” broadleaf tree forest type in central Asia. The plant population was widely regarded as a remnant of broadleaf Fig. 1 Similarity coefficients of Malus sieversii and M. pumila cultivars (types)
tree forest of warm temperate zone in Tertiary era[7]. M. sieversii was proposed to be the primary progenywww.jgenetgenomics.org
Xuesen Chen et al.: Genetic Diversity of Volatile Components in Xinjiang Wild Apple (Malus sieversii)
Table 3
177
Character impact odors in Malus sieversii and M. pumila cultivars
No. RT (min) 1
9.40
2
Compounds
Formula
Odor thresholds(μg/L)
Odor units Malus sieversii*
‘Golden Delicious’
‘Starking’ ‘Ralls’
‘Fuji’
Hexanal
C6H12O
64
3.57 (0.24-8.21)
5.34
6.28
7.35
7.47
10.43
2-methyl-ethyl butanoate
C7H14O2
18
0.41 (0-1.26)
0
0.04
0
3.39
3
12.34
(E)-2-hexenal
C6H10O
17
21.37 (3.35-38.98)
81.13
51.68
39.78
28.81
4
16.16
Hexyl acetate
C8H16O2
2
1.10 (0-11.28)
17.97
54.98
0.55
39.83
5
5.50
1-butanol
C4H9O
80
1.35 (0-7.22)
0
0
0.87
0.13
6
8.67
Ethyl butanoate
C6H12O2
20
3.12 (0.08-9.88)
0
0.35
0.31
0.88
7
13.14
1-hexanol
C6H14O
500
0.32 (0-1.44)
0.25
0.27
0.35
0.13
8
15.61
Ethyl hexanoate
C8H16O2
14
1.37 (0-23.04)
0.19
0.54
0.17
0
9
18.52
3-octen-1-ol
C8H16O
1.4
0.26 (0-2.29)
0
0
0
0
10
20.83
Ethyl octanoate
C10H20O2
5
2.59 (0-9.45)
0
0
0
0
11
25.77
Damascenone
C13H18O
0.05
9.58 (0-46.98)
0
0
0
0
12
5.55
Propyl acetate
C5H10O2
48
0
0
1.27
0
1.31
13
11.34
(Z)-3-hexenal
C6H10O
0.25
0
0
0
14
11.85
1-butanol,2-methyl-, acetate
C7H14O2
30
0.01 (0-0.18)
0
9.43
0
0
15
12.20
Pentyl acetate
C7H14O2
7.5
0
0
0
0
1.40
16
14.37
3-furanmethanol
C5H6O2
5
0.02 (0-0.51)
2.13
0
0
0
17
18.77
Benzene acetaldehyde
C8H8O
4
0
0
1.87
0
0
177.20
37.29
*Variation range in brackets
tor of the cultivated apple that played a crucial role in domestication of global cultivated apple. Therefore, further research on M. sieversii is of importance in apple breeding, genetics, evolution, and germplam conservation. Supported by National Nature Science Foundation in 1980s, Lin et al.[10] constructed field stations at Daxigou, Huocheng, and Jiaowutuohai in the Ily Valley, and investigated M. sieversii for almost a decade. The palynologic study on M. sieversii showed significant difference in P/E values between pollens from different sites. It was suggested that M. sieversii found in central Asia is advanced form, whereas M. sieversii found in Xinyuan is original form[9]. Li et al.[12] discovered that patterns of peroxidase isoenzymes in M. sieversii, M. orientalis Ugliz., M. sylvestris L., and M. pumila Borkh. Subsp. chinensis Li Y. N. were similar, which indicated the www.jgenetgenomics.org
homology in the four species. These authors supposed the former three species as the ancestor of cultivated apple. Using molecular analysis, Forte et al. also suggested that M. sieversii was probably the ancestor of cultivated apple[13,16,17]. In the present study, volatile composition in M. sieversii and in the controlled M. pumila cultivars were qualitatively and quantitatively measured using head space-solid phase microextraction and gas chromatography-mass spectrometry. Similarity coefficient concerning the types of volatiles was used to study the evolutionary process of apple. The result that the similarity coefficients were in accordance with the evolution of apple cultivars (form) indicated that the strategy with the volatile composition to study the relationship and evolution of apple species was methodically feasible. M. sieversii accessions mani-
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fested broad genetic variation and significant difference in total content, amount, and content of each chemical class, segregation ratios, and contents of main volatile compounds. Comparison of volatile compounds between M. sieversii and M. pumila cultivars ‘Ralls’, ‘Fuji’, ‘Golden Delicious’ showed that the common compounds whose number were larger than 5 with the contents over 0.04 mg/L simultaneously between M. sieversii and M. pumila belonged to esters, alcohols, aldehydes or ketones. This suggests that fundamental identity in main compounds of M. sieversii and M. pumila cultivars and This also supports the hypothesis “M. sieversii is probably the ancestor of M. pumila”. However, 48 compounds that were not detected
References 1
2
3
stract). Guo GN, Yan ZL, Zhang SN. Review and outlook of apple breeding in China. Journal of Fruit Science, 2003, 20(2): 127-134 (in Chinese with an English abstract).
longed to character impact odors. This shows that in
3. 2
Chen XS, Xin PG, Du XG, Yang CY, Wen JH. Effect of delicious and golden delicious in new apple cultivar breeding. Journal of Shandong Agricultural University, 1994, 25(2): 236-248 (in Chinese with an English ab-
3-furanmethanol, and benzene acetaldehyde) be-
hybridization is possible.
Xin PG , Chen XS. Application of aalls in apple breeding and culture. Journal of Fruit Science, 1993, 10(2): 107-112 (in Chinese with an English abstract).
them, six compositions (i.e., propyl acetate, (Z)-3-
the domestication of M. pumila, introgression of other Malus species, except for M. sieversii, by interspecies
Vol.34 No.2 2007
acted as character impact constituents. In awareness of the genetic diversity and individuality of compositions and content of volatiles in M. sieversii, the crossing breeding using M. sieversii with the species parent was not only benefit for flavor improvement of M. pumila cultivars but also facilitate to realize the objective of ‘utilization conservation’ of M. sieversii.
in M. sieversii were unique to M. pumila. Among hexenal, 2-methyl-1-butanol acetate, pentyl acetate,
Acta Genetica Sinica
4
Li YN. Researches of germplasm resources of Malus Mill. Beijing: China Agriculture Press, 2001, 20-23 (in Chinese
5
The relationship between composition of volatile compounds and conservation of M. sieversii
with an English abstract). Gayle MV, Christopher MR, Reilley AA, Adam DH. Ex situ conservation of vegetatively propagated species: Development of a seed-based core collection for Malus sieversii. Journal of American Society of Horticulture Science, 2005, 130(2): 203-210.
Volatile components are important factors that determine the fruits flavor quality. Recent research has shown that the whole aroma of apple not only directly relates to the content of volatile compounds but also directly relates to the number of volatile compounds. Sensory interaction leads to a strong sensory impact on an apple cultivar, which contains more types of volatile compounds[18]. The present study discovered that M. sieversii showed great genetic diversity in types and content of volatile compounds. Ninety unique compounds were detected in M. sieversii and seven compounds including γ-buty-
rolactone, methyl-γ-butyrolactone, acetals, 1-butanol, ethyl butanoate, ethyl octanoate, and damascenone
6
Forsline PL, Aldwinckle HS. Evaluation of Malus sieversii seedling populations for disease resistance and horticultural traits. Acta Horiculturae, 2004, 663(2): 529-534.
7
Zhang XS. On the eco-geographical characters and the problems of classification of the wild fruit-tree forest in the Ily valley of sinkiang. Acta Botanica Sinica, 1973, 15(2): 239-253 (in Chinese with an English abstract).
8
Zheng DS, Yang QW. In situ conservation site construction of agricultural wild plants in China. Journal of Plant Genetic Resources, 2004, 5(4): 386-388 (in Chinese with an
9
English abstract). Yang XH, Li YN, Lin PJ. Study on the pollen morphology of Malus sieversii in China and its origin and evolution. Journal of Southwest Agricultural University, 1992, 12(1): 45-50 (in Chinese with an English abstract).
10 Lin PJ, Cui NR. Wild fruit forests resources in Tianshan www.jgenetgenomics.org
Xuesen Chen et al.: Sequence Variation and Molecular Evolution of Hormone-Sensitive Lipase Genes in Species of Bovidae
Mountains: comprehensive research on wild forests in Lly, Xinjiang, China. Beijing: China Forest Press, 2000, 8-9
1-6 (in Chinese with an English abstract). 15 Chen MX, Chen XS, Feng BC. GC-MS analysis of fruit aroma components of two apricot cultivars. Acta Hor-
(in Chinese with an English abstract). 11 Liu J, Zhou QH, Sun HW, Qiu ZL, Wang YW, Sun ZX.
ticulturae Sinica, 2004, 31(5): 663-665 (in Chinese with
Study on the phenotype biodiversity of Xinjiang wild apples (Malus sieversii). Journal of Fruit Science, 2004,
179
an English abstract). 16 Zhang L, Cai YM, ZhuGe Q, Lou LH, Zou HY, Huang
21(4): 285-288 (in Chinese with an English abstract).
MR, Wang MX. Analysis of the inter-species relation-
12 Li YN, Li XL. Studies on the pox isoenzyme banding pat-
ships on Lycoris (Amaryllidaceae) by use of RAPD. Acta
terns of the genus Malus in the world. Journal of South-
Genetica Sinica, 2002, 29(10): 915-921 (in Chinese with
west Agricultural University, 1995, 17(5): 371-377 (in
an English abstract).
Chinese with an English abstract).
17 Zhang L, Cai YM, ZhuGe Q, Zou HY, Huang MR. Se-
13 Forte AV, Ignatov AN. Phylogeny of the Malus (apple tree)
quence of the ITS region of nuclear ribosomal DNA
species, inferred from the morphological traits and mo-
(nrDNA) in Xinjiang Wild Dianthus and its phylogenetic
lecular DNA analysis. Russian Journal of Genetics, 2002,
relationship. Acta Genetica Sinica, 2002, 29 (6): 549-554
38(10): 1150-1160. 14 Wang LP, Xu Y, Zhao GA, Chen J, Wang LP, Liu YM, Li JM. Rapid determination of components in apple wine by head space solid-phase microextraction (HS-SPME). Journal of Wuxi Light Industry University, 2003, 22(1):
(in Chinese with an English abstract). 18 Johannes HFB, Hendrik NJK, Jacques PR, Estanislau DB,
Alphons GJV, Jan HAK. Sensory evaluation of character impact components in an apple model mixture. Chemical Senses, 2002, 27: 485-494.
新疆野苹果挥发性化合物组分的遗传多样性研究 陈学森, 冯 涛, 张艳敏, 何天明, 冯建荣, 张春雨 山东农业大学果树生物学实验室，泰安 271018
摘 要：以中国新疆伊犁地区巩留县莫合镇的新疆野苹果（Malus sieversii (Lebed.) Roem.）30 个实生株系及国光、元帅、富 士、金冠等苹果（Malus pumila Mill.）品种的成熟果实为试材，采用顶空固相微萃取和气相色谱质谱联用技术，分析香气 成分，旨在为探讨新疆野苹果与栽培苹果的亲缘关系及为新疆野苹果资源保护利用提供基本资料。结果表明，①根据新疆 野苹果与栽培苹果共有挥发性化合物种类数计算相似系数，其大小与苹果品种（类型）的演化历史相吻合；②新疆野苹果 各实生株系挥发性化合物总含量、各类挥发性化合物种类数及其含量以及主要挥发性化合物分离比率与含量等存在广泛的 遗传变异，参试的 30 个实生株系间差异明显，遗传多样性极为丰富；进一步与 4 个栽培苹果品种挥发性化合物组分进行比 较，结果发现，新疆野苹果与栽培苹果品种化合物种类数在 5 以上、含量在 0.04 mg/L 的化合物均属于酯类、醇类、醛类 和酮类，说明新疆野苹果与栽培苹果的主要挥发性化合物组分基本一致，上述研究结果支持“新疆野苹果可能是栽培苹果的 祖先种”的结论，但有 48 种成分为栽培苹果特有的挥发性化合物成分，在新疆野苹果检测不到，其中乙酸丙酯及苯乙醛等 6 种成分为特征香气成分。因此，栽培苹果可能是杂种起源；③从新疆野苹果 30 个实生株系中共检测到包括酯类、醇类、 酮类、醛类、酸类、苯衍生物、杂环类、萜类、烃类、缩醛类和内酯类等 11 类化合物 177 种，其中缩醛类和内酯类化合物 在栽培苹果品种中没有检测到，90 种成分为新疆野苹果特有成分，1-丁醇及大马酮等 7 种成分为香气值大于 1 的特征香气 成分。因此，新疆野苹果作为栽培苹果遗传改良的珍贵资源，进行“利用保存”的潜力很大。 关键词：新疆野苹果；栽培苹果；挥发性化合物；遗传多样性 作者简介：陈学森（1958-），男，山东临沭人，博士，教授，博士生导师，研究方向：果树种质资源与遗传育种。 E-mail:
[email protected] www.jgenetgenomics.org
