全 文 ：High-frequency Plant Regeneration from Leaf
Cultures of Moringa oleifera Lam: a Multipurpose
Plant
Guocong LI, Qiyan ZHOU, Xin LI*
Guangxi Zhuang Autonomous Region, Qinzhou Environmental Monitoring Centre, Qinzhou 535000, China
National Natural Science Foundation of China (21165008).
*Corresponding author. E-mail: 283223173@qq.com
Received: April 1, 2016 Accepted: May 17, 2016A
Agricultural Science & Technology, 2016, 17(6): 1318-1321, 1358
Copyright訫 2016, Information Institute of HAAS. All rights reserved Molecular Biology and Tissue Culture
M oringa oleifera Lam (syn. M.ptreygosperma Gaertn.) isone of the best known and
most widely distributed and natural-
ized species of a monogeneric fam-
ily Moringaceae [1-2]. This tree grows in
any tropical and subtropical country
with peculiar environmental features,
namely, dry to moist tropical or sub-
tropical clime, with annual precipitation
of 760 to 2 500 mm (it requires less
than 800 mm irrigation) and tempera-
ture between 18 and 28 ℃ [3]. It grows
in any soil type, but heavy clay and
waterlogged, with pH between 4.5 and
8, at an altitude up to 2 000 m[4-5].
Moringa oleifera is an important
food commodity which has had enor-
mous attention as the “natural nutri-
tion of the tropics”. The leaves, fruit,
flowers and immature pods of this
tree are used as a highly nutritive veg-
etable in many countries, particularly
in India, Pakistan, Philippines, Hawaii
and many parts of Africa [6-8]. Moringa
leaves have been reported to be a
rich source of β-carotene, protein, vi-
tamin C, calcium and potassium and
act as a good source of natural an-
tioxidants; and thus enhance the
shelf-life of fat containing foods due to
the presence of various types of an-
tioxidant compounds such as ascorbic
acid, flavonoids, phenolics and
carotenoids[9-12]. In the Philippines, it is
known as “mother’s best friend” be-
cause of its utilization to increase
woman’s milk production and is so-
metimes prescribed for anemia[10, 12-14].
Since it is a multipurpose plant
and perennial in nature, cultivation of
M. oleifera assumes utmost impor-
tance to meet the large-scale demand
and ensure continuous supply of the
elite material. Traditional propagation
through stem cuttings is possible, but
low seed yield and easy uprooting of
established plants on poor and
Abstract [Objective] The aim was to optimized and established the regeneration
system of Moringa oleifera and provided foundation for its rapid propagation and fur-
ther research. [Method] Tissue culture techniques and large scale micropropagation
of M. oleifera were studied in this paper． By means of a series of experiments, we
used the leaf of aseptic seedling from M. oleifera as explants to optimize and es-
tablish the regeneration system cultured in vitro by means of direct organogenesis.
[Result] It was observed that using the fresh shelled M. oleifera seeds with 0.1%
mercuric chloride for 6 minutes could reach the best disinfection effect. The seed
germination rate was 85% . The leaf could produce cluster buds well using the
medium with MS+2.0 mg/L 6-BA+0.05 mg/L NAA, while the best proliferation condi-
tion was under MS+6-BA 1.0 mg/L+KT 0.1 mg/L+2, 4-D 2.0 mg/L+NAA 0.05 mg/L.
The best rooting induction culture medium was MS+0.5 mg/L IBA, with the rooting
rate as 100%. [Conclusion] This protocol might find use in mass production of true-
to-type plants and in production of transgenic plants through Agrobacterium/biolistic-
mediated transformation.
Key words Moringa oleifera; Tissue culture; Proliferation; Rooting; Optimization
辣木组织培养及高频率植株再
生
李国葱，周启艳，李欣* （广西壮族自治区钦
州市环境保护监测站，广西钦州 535000）
摘 要 [目的]为辣木组培快繁及规模化育苗
提供技术参考。 [方法]以优选辣木无菌苗的真
叶为材料，采用组织培养技术对其诱导、增殖、
生根等问题进行研究。 [结果]新鲜种子去壳后
用 0.1%升汞处理 6 min 的消毒效果最好，成功
率达 85%。 辣木无菌苗真叶不定芽诱导及增殖
最佳培养基分别为 ：MS+6-BA 1.0 mg/L+KT
0.1 mg/L+2, 4-D 2.0 mg/L+NAA 0.05 mg/L、
MS+0.5 mg/L 6-BA+0.03 mg/L NAA，丛生芽明
显 。 辣木组培苗最佳生根培养基为 MS+0.5
mg/L IBA，生根率可达 100%，炼苗移栽后成活
率达 98%以上。 [结论]辣木外植体的离体培养
表明，不同的激素组合及其浓度对辣木外植体
诱导效果差别大， 各种激素经过优化配合，可
以获得更高的诱导率，本研究为辣木的无菌快
繁奠定应用基础，并为辣木基因工程育种等研
究提供技术参考。
关键词 辣木 ; 组织培养 ; 增殖 ; 生根 ; 体系
优化
基 金 项 目 国 家 自 然 科 学 基 金 项 目
(21165008)。
作者简介 李国葱（1968-），女，广西钦州人 ，
助理工程师， 从事环境监测工作研究，E-mail：
zqypwh@126.com。 *通讯作者，硕士，从事环境
监测工作研究，E-mail：35538556@qq.com。
收稿日期 2016-04-01
修回日期 2016-05-17
DOI:10.16175/j.cnki.1009-4229.2016.06.009
Agricultural Science & Technology2016
marginal soils hamper the practical ut-
ility of this propagation method. Con-
sequently, a rapid clonal multiplication
technique such as micropropagation
could be applied to M. oleifera, which
will supplement conventional culturing
methods and lead to rapid improve-
ment of the tree. Although micropropa-
gation in M. oleifera has been at-
tempted using explants of cotyledon,
the presence of intermediary callus or
callus-mediated regeneration is least
desired for the production of true-to-
type plants[15].
In this study, we investigated in
vitro adventitious shoot-bud induction
competency of leaf from M. oleifera. In
the process, we established an effi-
cient protocol for high-frequency direct
regeneration of plantlets from leaf of
M. oleifera.
Materials and Methods
Plant materials
M. oleifera seeds were collected
from India, The decoated seeds were
soaked in distilled water for 48 h at
room temperature. After soaking, de-
coated seeds were surface sterilized
with 1% (w/v) 84 disinfectant (shine
life, China) for 30 min and subsequent-
ly rinsed three times with sterile dis-
tilled water. After treating with disinfec-
tant, the seeds were treated with 0.1%
(w/v) mercuric chloride for 6 min fol-
lowed by sterile distilled water rins-
ing. The material was blotted dry on
sterile filter paper, and then the dis-
infected seeds were inoculated on
Murashige and Skoog [16] basal medi-
um without any growth regulators at
25±2 ℃ and maintained under a 12 h
photoperiod of 90 μmol/m2·s irradi-
ance provided by cool-white fluores-
cent tubes with a relative humidity of
70%.
Adventitious shoot induction
The young leaves (2 -3 mm in
length) were dissected from 3-week-
old in vitro cultured seedlings and cul-
tured on MS basal medium supple-
mented with different combinations of
6-benzyladenine (6-BA) and 2,4-
dichlorophenoxyacetic acid (2, 4-D)
and 6-Furfurylaminopurine (KT) and α-
naphthalene acetic acid (NAA) for ad-
ventitious shoot induction (Table 1). All
medium was supplemented with 3.0%
(w/v) sucrose and 0.7% (w/v) agar
(Beijing Dingguo Changsheng
Biotechnology Co., Ltd.). The medium
pH was adjusted to 5.8 with 1.0 M
NaOH prior to autoclaving at 121℃ for
20 min. The medium was aliquoted in-
to sterile 250 ml glass culture vessels
(25 ml each). There were 10 explants
per treatment, and three replications
were conducted. Culture conditions
were the same as described above.
Adventitious shoot induction frequen-
cy, defined as the percentage of ex-
plants with adventitious shoots, as well
as the number of shoot per explant
was recorded after 4 weeks of culture.
All plant growth regulators were pur-
chased from Beijing Dingguo Chang-
sheng Biotechnology Corporation,
Limited.
Shoot proliferation
Micro-shoots were sub-cultured
in hormone-free MS basal medium
for 1 week to eliminate any carry-
over effects of the basic hormone [17].
For shoot proliferation, micro-shoots
approximately 1 cm in length were
sub-cultured in MS basal medium
supplemented with 6-BA and NAA at
different concentrations (Table 2). For
each replicate, 25 ml of the medium
was dispensed into 250 ml culture
vessels. Each treatment contained 10
micro-shoots and was repeated three
times. Culture conditions were the
same as described above. After 20
days of culture, number of available
shoots and micro-shoot growing status
were recorded.
Rooting of adventitious shoots
For in vitro root formation, micro-
shoots about 2 -3 cm in length were
transferred to half-strength MS basal
medium supplemented with 0.0, 0.1,
0.3, 0.5, 0.7 and 1.0 mg/L IBA. Three
replications with five explants per
treatment were conducted. Micro-
shoots on root induction medium were
cultured in the dark for 2 days at 25 ± 2
℃ prior to being cultured under a 12 h
photoperiod (90 μmol/(m2·s)). After 15
days of culture, the root formation fre-
quency, root number, root length, and
number of lateral roots were deter-
mined.
Acclimatization of in vitro-rooted
plantlets
Plantlets in the culture vessels
were transferred from the culture room
to the greenhouse for 3-4 days. Plan-
tlets with well-developed shoots and
roots were then removed from the a-
gar medium and potted in 10-cm plas-
tic pots containing a soil-less substrate
(peat moss and loam in a 2:1 volumet-
ric ratio). Agar was removed from the
roots thoroughly prior to transplanting.
The pots were covered with polythene
films to provide a high relative humidi-
ty, and the films were gradually
opened after 1 week in the green-
house to allow plantlets to acclimatize
Table 1 Experiment design of different level of plant growth regulators
No.
Plant growth regulators//mg/L
6-BA KT 2，4-D NAA
LM-1 0.0 0.05 0.5 0.0
LM-2 0.0 0.1 1.0 0.05
LM-3 0.0 0.2 2.0 0.1
LM-4 1.0 0.05 0.5 0.0
LM-5 1.0 0.1 2.0 0.05
LM-6 1.0 0.2 1.0 0.1
LM-7 2.0 0.05 1.0 0.0
LM-8 2.0 0.1 0.5 0.05
LM-9 2.0 0.2 2.0 0.1
LM-10 3.0 0.05 1.0 0.0
LM-11 3.0 0.1 2.0 0.05
LM-12 3.0 0.2 0.5 0.1
LM-13 4.0 0.05 2.0 0.0
LM-14 4.0 0.1 0.5 0.05
LM-15 4.0 0.2 1.0 0.1
CK 0.0 0.0 0.0 0.0
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Agricultural Science & Technology 2016
to ambient conditions. Plants were
watered every 2-3 days until the films
were fully opened, and then watered
as required
Results
Adventitious shoot formation
For adventitious shoot induction,
young leaves from in vitro-germinated
plants were used as explants. Adven-
titious shoots were mainly developed
from the basipetal portion around the
cut ends of young leaves after 4 weeks
of culture, and its development contin-
ued until the fifth week, when the re-
sults were recorded. Significant differ-
ences in the rates of callus were ob-
served in leaf explants. Highest rates
of callus (66.59%) were obtained (LM-
7) at the concentrations of 6-BA
(2.0mg/L) in combination with KT (0.05
mg/L) and 2, 4-D (1.0 mg/L), which
yielded 1.27 shoots per young leaf ex-
plant. Following the treatments (LM-5)
of 6-BA (1.0 mg/L) combined with KT
(0.1 mg/L) and 2, 4-D (2.0 mg/L) and
NAA (0.05 mg/L), the treatments (LM-
11) of 6-BA (3.0 mg/L) combined with
KT (0.1 mg/L) and 2, 4-D (2.0 mg/L)
and NAA (0.05 mg/L). Their callus for-
mation rate was 54.17% and 56.73%,
respectively. Although LM-7 combina-
tion could get the highest rate of callus,
LM-5 and LM-11 combinations could
induce more adventitious shoots,
yielded 3.81 and 2.43 shoots per
young leaf explant, respectively, when
compare with LM-7. Moreover, when
other regulators are the same, with the
increasing concentrations of 6-BA, the
callus formation frequency increased
gradually in all treatments, but the for-
mation rate of adventitious shoot was
decreased. In general, poorer
organogenesis was the result of low
(0.0 mg/L) or high (2.0 mg/L) 6-BA
concentrations. Therefore, the best
adventitious shoot formation culture
medium found in this study was LM-5:
MS + 6-BA 1.0 mg/L + KT 0.1 mg/L + 2,
4-D 2.0 mg/L + NAA 0.05mg/L (Fig.1A).
Shoot proliferation
Micro-shoot proliferation was
achieved by all treatments (Table 2).
However, the greatest proliferation oc-
curred in MS medium containing
0.5mg/L 6-BA and 0.03 mg/L NAA,
which stimulated M. oleifera shoot el-
ongation and proliferation most effec-
tively, and its multiplication coefficient
was 3.4 (Table 2). The shoots were
robust and healthy, exhibiting a dark
green color (Fig.1B). Shoots grew
rapidly in this medium and nodal seg-
ments were sub-cultured every 5 days.
Micro-propagation is amenable to in
vitro culture on M. oleifera. In hor-
mone-free MS medium, the number of
available shoots per explant was 0.4.
The number of adventitious buds in-
creased in all treatments with increas-
ing concentrations of 6-BA and in
combination with 0.03 mg/L NAA; how-
ever, the number of available shoots
per explant was decreased when NAA
concentrations exceeded 0.05 mg/L
(Table 2). Moreover, shoots produced
at higher concentrations of 6-BA were
pale green, and then turn into brown
and withered.
In vitro rooting
The micro-shoots were trans-
ferred to root induction medium to in-
duce rooting (Fig.1C), and developed
directly from the base of the stems
with minimal callus formation after 20
days (Fig. 1D). Exogenous auxin had
a significant effect on root induction
rates and the number of roots, com-
pared with the control treatment (hor-
mone-free medium) (Table 3). Exclud-
ing the control treatment, the root in-
duction rate was more than 85.7% and
at least 6.43 roots were formed. The
highest rooting percentage (100) was
induced by 0.5 mg/L IBA, a high num-
ber of roots (7.94) was obtained. As
shown in table 3, IBA had a positive
effect on root length and the number
of lateral roots. In culture medium
containing 0.5 mg/L IBA, both root
length and the number of lateral roots
were the best. Therefore, regarding in
vitro root formation of M. oleifera, the
best culture medium was MS + 0.5
mg/L IBA.
Ex vitro transplanting
After 30 days of culture on root-
Table 2 Effect of 6-BA combined with NAA on shoot proliferation
No.
ρ//mg/L Multiplication
coefficient No.
ρ//mg/L Multiplication
coefficient6-BA NAA 6-BA NAA
ZJ-1 0.1 0.01 1.2 ZJ-9 0.5 0.01 2.6
ZJ-2 0.1 0.03 1.4 ZJ-10 0.5 0.03 3.4
ZJ-3 0.1 0.05 1.4 ZJ-11 0.5 0.05 2.7
ZJ-4 0.1 0.1 1.6 ZJ-12 0.5 0.1 3.0
ZJ-5 0.3 0.01 2.3 ZJ-13 1 0.01 1.1
ZJ-6 0.3 0.03 2.4 ZJ-14 1 0.03 1.4
ZJ-7 0.3 0.05 2.6 ZJ-15 1 0.05 1.7
ZJ-8 0.3 0.1 2.7 ZJ-16 1 0.1 1.9
Table 3 Effect of IBA on In vitro root induction
No. IBA//mg/L Root rate//% Root number Root length//cm Lateral roots number Root status
1 0 65.3 4.11 1.08 0.02 Slender
2 0.1 85.7 6.43 1.25 1.74 Slender
3 0.3 92.1 8.01 1.17 1.64 Short, thick
4 0.5 100 7.94 1.34 0.96 Shorter, thicker
5 0.7 97.4 7.54 1.29 1.42 Dense and shorter
6 1.0 91.7 6.87 0.94 1.70 Dense and shorter
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Agricultural Science & Technology2016
Fig.1 Adventitious shoot-bud induction and plant regeneration from leaf cultures
of M. oleifera
induction medium, 80% rooting was
observed without any intermediary
callus growth. The well-developed
plantlets were washed with sterile wa-
ter to remove traces of agar from the
roots and dipped in 0.1% (w/v) of a
broad-spectrum fungicide solution (84
disinfectant) for 10 min and transferred
to the potting mixture containing 1:1
ratio of cocopeat:garden soil in 3 inch
diameter pots. Primary hardening took
place in 3-4 weeks under high-humid-
ity conditions created by covering pots
with polythene bags. The well accli-
matized plants were further transferred
to the field with more than 98% sur-
vival rate.
Discussion and Conclusion
Explant treatment with different
plant growth regulators is a common
technique generally employed for
getting maximum results[18]. Of the dif-
ferent cytokinins tested separately,
none of them produced shoots from
the young leaf explants. In the present
study, 6-BA combined with KT and 2,
4-D and NAA were able to induce cel-
lus in vitro regeneration. These results
were similar to those of previous stud-
ies on M. oleifera [15]. In addition, the
other important factors influencing M.
oleifera adventitious shoot induction in-
cluded the type and nature of the ex-
plants. In vitro young leaves (20 days
old) as explants provided higher ad-
ventitious shoot induction frequency
than did mature one. Additionally, we
found that moderate concentration of
6-BA in combination with the same
concentrations of KT and 2, 4-D and
NAA was superior to lower or higher
concentration of 6-BA in terms of M.
oleifera adventitious shoot formation,
which suggested that 6-BA may play
a main factor in vitro on young leaf.
It is typically difficult for roots to de-
velop from in vitro woody shoots,
while in this study, in vitro M. oleifera
shoots rooted in medium at a high
frequency (85.7%-100%). This is simi-
lar to Caesalpinia bonduc[19], Carica pa-
paya [20], Santalum album [21], Gardenia
latifolia [22], and Caesalpinia bonduc [23].
All rooting percentages were at least
70% , and some reached 100% . The
significant differences in rooting per-
centages among woody plants may be
due to different levels of endogenous
hormones in the explants.
In conclusion, the availability of
highly efficient regeneration protocol
using young leaf is highly desirable for
rapid mass clonal propagation, con-
servation strategies, germplasm ex-
change, producing phytomedicines. It
is also desirable for Agrobacterium
tumefaciens-mediated genetic trans-
formation to introduce useful genes
such as virus, insect, and cold resis-
tance genes for increasing resistance
of this important species.
References
[1] NADKARNI A K. Indian Materia Medica
[M]. Popular Prakashan: Bombay, 1976,
pp: 810-816.
[2] RAMACHANDRAN C, PETER K V,
GOPALAKRISHNAN P K. Drum-stick
(Moringa oleifera): a multipurpose Indi-
an vegetable [J]. Econ Bot, 1980, 34:
276-283.
[3] MORTON J F. The horseradish tree,
Moringa pterigosperma (Moringaceae).
A boon to arid lands [J]. Econ Bot,
1991, 45: 318–333.
[4] QAISER M. Moringaceae. In Flora of
West Pakistan, Nasir E, Ali SI (eds) [M].
No.38. University of Karachi Press:
Karachi, 1973, pp: 1–4.
[5] PALADA M C, CHANGL L C. Suggest-
ed cultural practices for Moringa. Inter-
national Cooperators’ Guide AVRDC
[J]. AVRDC pub#03 -545 www.avrdc.
org. 2003.
[6] ANWAR F, BHANGER M I. Analytical
characterization of Moringa oleifera
seed oil grown in temperate regions of
Pakistan [J]. J Agric Food Chem, 2003,
51: 6558-6563.
[7] ANWAR F, ASHRAF M, BHANGER M I.
Inter provenance variation in the com-
position of Moringa oleifera oil seeds
from Pakistan [J]. J Am Oil Chem Soc,
2005, 82: 45-51.
[8] BERKOVICH L, EARON G, RON I, et
al. Moringa Oleifera aqueous leaf ex-
tract down-regulates nuclear factor-
kappa B and increases cytotoxic effect
of chemotherapy in pancreatic cancer
cells [J]. BMC Complem Altern M, 2013,
13: 212.
[9] BAKRE A G, ADERIBIGBE A O, ADE-
MOWO O G. Studies on neuropharma-
cological profile of ethanol extract of
Moringa oleifera leaves in mice [J]. J
Ethnopharmacol, 2013,149: 783-789.
[10] SIDDHURAJU P, BECKER K. Antioxi-
dant properties of various solvent ex-
tracts of total phenolic constituents
from three different agro-climatic ori-
gins of drumstick tree (Moringa
oleifera Lam.) [J]. J Agric Food Chem,
2003, 15: 2144-2155.
[11] SREELATHA S, JEYACHITRA A, PA-
DMA P R. Antiproliferation and induc-
tion of apoptosis by Moringa oleifera
leaf extract on human cancer cells [J].
Food Chem Toxicol, 2011, 49: 1270-
1275.
[12] ESTRELLA M C P, MANTARING J B
V, DAVID G Z. A double blind, ran-
domised controlled trial on the use of
malunggay (Moringa oleifera) for aug-
mentation of the volume of breast milk
among non-nursing mothers of
preterm infants [J]. Philipp J Pediatr,
(Continued on page 1358)
1321
Agricultural Science & Technology 2016
Responsible editor: Xiaoxue WANG Responsible proofreader: Xiaoyan WU
2000, 49: 3-6.
[13] HANNAN M A, KANG J Y, MOHIB-
BULLAH M, et al. Moringa oleifera with
promising neuronal survival and neu-
rite outgrowth promoting potentials [J].
J Ethnopharmacol, 2014, 152: 142 -
150.
[14] MOYO B, OYEDEMI S, MASIKA P J,
et al. Polyphenolic content and antiox-
idant properties of Moringa oleifera
leaf extracts and enzymatic activity of
liver from goats supplemented with
Moringa oleifera leaves/sun flower
seed cake [J]. Meat Sci, 2012, 91:
441-447.
[15] ZHANG Q. The study on the tissue
culture system and determine the ef-
fective content of Moringa oleifera [D].
Fujian Agriculture and Forestry Uni-
versity, 2013.
[16] MURASHIGE T, SKOOG F. A revised
medium for rapid growth and bioas-
says with tobacco tissue culture [J].
Physiol Plant, 1962, 15: 473-497.
[17] SHIBLI R A, SHATNAWI M, ABU-EIN,
et al. Somatic embryogenesis and
plant recovery from callus of ‘Nabali’
olive (Olea europea L.) [J]. Sci Hortic
(Amsterdam), 2001, 88: 243-256.
[18] D’ONOFRIO C, MORINI S. Somatic
embryo, adventitious root and shoot
regeneration in vitro grown quince
leaves as influenced by treatments of
different length with growth regulators
[J]. Sci Hortic, 2006, 107: 194-199.
[19] CHERUVATHUR M K, BRITTO J,
THOMAS T D. Pulvinus: an ideal ex-
plant for plant regeneration in Cae-
salpinia bonduc (L.) Roxb., an impor-
tant ethno medicinal woody climber [J].
Acta Physiol Plant, 2012, 34: 693-699.
[20] ROY P K, ROY S K, HAKIM M L.
Propagation of papaya (Carica papaya
L.) cv. shahi through in vitro culture [J].
Bangladesh J Bot, 2012, 41: 191-195.
[21] SINGH C K, RAJ S R, PATIL V R, et
al. Plant regeneration from leaf ex-
plants of mature sandalwood (San-
talum album L.) trees under in vitro
conditions [J]. In Vitro Cell Dev-Pl,
2013, 49: 216-222.
[22] REDDY Y M, SARITHA K V. In vitro
clonal propagation of Gardenia latifolia
Ait.: A toy making woody tree [J]. Ag-
rofor Syst, 2013, 87: 591-598.
[23] KUMAR S, KRISHNA V, VENKATE-
SH, et al. Direct and indirect method of
plant regeneration from root explants
of Caesalpinia bonduc (L.) Roxb.: A
threatened medicinal plant of Western
Ghats [J]. Indian J Exp Biol, 2012, 50:
910-917.
(Continued from page 1321)
Responsible editor: Yingzhi GUANG Responsible proofreader: Xiaoyan WU
2002, 29(4): 6-8.
[2] XIAO XC (肖兴翠). Study on the effect
of different silvicultural measures on
growth and nutrient cycling of slash pine
(不同营林措施对湿地松林分生长及养
分 循 环 的 影 响 研 究 ) [D]. Changsha:
South Central Forestry S&T university
(长沙: 中南林业科技大学), 2013.
[3] XIAO XC (肖兴翠 ), LI ZH (李志辉 ),
TANG ZJ (唐作钧 ), et al. Effects of
stand density on biomass and produc-
tivity of Pinus elliottii (林分密度对湿地松
生物了及生产力的影响 ) [J]. Journal of
Central South Forestry University (中南
林业科技大学学报), 2011, 31 (3): 123-
128.
[4] ZHUANG WY (庄伟瑛), ZHANG YY (张
玉英), ZOU YX (邹元熹). Selection for
high-resin yield of slash pine and analy-
sis of factors concerned (高产脂湿地松
选择和相关因子的分析)[J]. Acta Agri-
culturae Universitis Jiangxiensis (江西
农业大学学报), 2007, 29(1): 55-60,65.
[5] GONG M (宫铭 ). Studies on the fast
propagation in vitro of high-yield resin
slash pine (高产脂湿地松组培快繁技术
研究)[D]. Nanchang: Jiangxi Agricultural
University (南昌:江西农业大学), 2013.
[6] LI YJ (李彦杰). Genetic analysis and ev-
aluation of slash pine families on resin
production related traits (湿地松家系产
脂性状遗传分析与评价技术研究 ) [D].
Beijing: Chinese Academy of Forestry
Sciences (北京: 中国林业科学研究院),
2013.
[7] YANG YJ (杨予静 ), LI CX (李昌晓 ),
ZHANG Y (张晔), et al. Effects of sub-
mergence and drought alternation on
nutrient contents in the soil growing
slash pine (Pinus elliottii) seedlings (水
淹-干旱交替胁迫对湿地松幼苗盆栽土
壤营养元素含量的影响 ) [J]. Scientia
Silvae Sinicae (林业科学), 2013, 49(2):
61-70.
[8] WANG ZX (王振夏), WEI H (魏虹), LI
CX (李昌晓), et al. Effect of soil mois-
ture variations on photosynthetic char-
acteristics of slash pine seedlings (土壤
水分交替变换对湿地松幼苗光合特性的
影响 ) [J]. Acta Botanica Boreali -Occi-
dentalia Sinica （西北植物学报 ), 2012,
32(5): 0980-0987.
[9] ZHOU J (周珺), WEI H (魏虹), LYU (吕
茜), et al. Effects of soil water regime on
leaf photosynthetic characteristics of
slash pine (Pinus elliottii Engelm.)
seedlings (土壤水分对湿地松幼苗光合
特征的影响 ) [J]. Chinese Journal of E-
cology (生态学杂志), 2012, 31 (1):30-
37.
[10] NIU T (牛涛), WANG YK (汪有科), WU
PT (吴普特), et al. Effects of difference
soil moisture conserving ways on soil
moisture, soil temperature and growth
of jujube (不同保墒措施对枣园土壤温、
湿度及枣树生长特征的影响)[J]. Jour-
nal of Irrigation and Drainage (灌溉排
水学报), 2008, 27(1): 35-38.
[11] HUANG J (黄俊 ), ZHANG NU (张弩 ),
MIN ZP (闵泽萍). Study on the effects
of different ground mulching materials
on the quality of early-maturing Citrus
unshiu and soil temperature and mois-
ture of Citrus unshiu orchard (不同地
面覆盖材料对早熟温州蜜柑果实品质
及桔园土壤温度和水分的影响效应研
究)[J]. South China Fruits (中国南方果
树),2010,39(3):15-17.
[12] DAO JM (刀静梅), LIU SC (刘少春),
ZHANG YB (张跃彬), et al. Effects of
whole film mulching on character of
non-irrigation sugarcane and soil tem-
perature and water (地膜全覆盖对旱地
甘蔗性状及土壤温湿度的影响 ) [J].
Sugar Crops of China (中国糖科 ),
2015, 37(1): 22-23
[13] LI JX (李进学), ZHOU DG (周东果),
ZHU YH (朱红业), et al. Effects of dif-
ferent cover methods on soil tempera-
ture-humidity changes, growth and
yield in lemon orchard (不同覆盖对柠
檬园土壤温湿度及其生长结果的影响)
[J]. Southwest China Journal of Agri-
cultural Sciences （西南农业学报 ）,
2010,23(2):487-491.
[14] YANG XK (杨相昆), WEI JJ (魏建军 ),
ZHANG ZQ (张占琴 ), et al. Effect of
degradable films on cotton soft tem-
perature and humidity (可降解地膜对
棉田土壤温湿度的影响)[J]. Southwest
China Journal of Agricultural Sciences
(西南农业学报 ), 2015, 28 (4): 1736-
1741.
[15] LIU P (刘平), MA LY (马履一), HAO
YR (郝亦荣). Effect of eco-mat on soil
temperature and water content and
weed of afforestation in river-beach (生
态垫对河滩造林地土壤温湿度和杂草
的影响 )[J]. Science of Soil and Water
Conservation (中国水土保持科学 ),
2005, 3(1): 77-81.
[16] WANG KX (王开喜). Effect of different
soil moisture conserving ways on soil
moisture and soil temperature in the
jujube slope of loess hilly region (不同
保墒措施对黄土丘陵区坡地枣园土壤
温、湿度的影响)[J]. China Rural Water
and Hydropower (中国农村水利水电),
2011, 7: 21-23.
[17] SUI YH (隋益虎), ZHAO X (赵勋). Ef-
fects of different mulching methods on
soil temperature and humidity and
early maturing culture of purple cab-
bage (不同覆盖方式的土壤温、湿度变
化及对紫甘蓝早熟栽培的影响 ) [J].
Soils (土壤), 2000,3:155-159.
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