The type,position and age effect on the cutting reproduction of Picea crassifolia and its rooting mechanism in the Qilian Mountains

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ORIGINAL PAPER

The type,position and age effect on the cutting reproduction of Picea crassifolia and its rooting mechanism in the Qilian Mountains

Jianjun Kang1,2•Wenzhi Zhao1•Ming Zhao2•Guangyu Li2•Jiqiang Zhang2•Zhisheng Zhang3

©Northeast Forestry University and Springer-Verlag Berlin Heidelberg 2015

Picea crassifoliaKom,a perennial arbor species is recognized as one of the most adaptable plants found to date in Qilian Mountains.To explore the cutting reproduction technology ofP.crassifoliaand reveal its rooting mechanism,cuttings ofP.crassifoliawith different cutting types(softwood,hardwood and root),positions (top,upper,middle and bottom)and ages(7,10,15,20, 25 year-old)were cultivated in a f i eld experiment.Oneyear old softwood and hardwood cuttings were collected from 7-,10-,15-,20-,and 25 year-old healthy ortets to analyze the changes from endogenous hormones and organic nutrients.Results indicate that the softwood cuttings(0.5–1.0 cm in diameter)from upper branches of 15 year-old ortets shows better growth performance by improving rooting indexes,including a signif i cant increase in rooting rate and a decrease in basal rot rate.Concomitantly,increasing rooting quantity and root length also increased.It is noteworthy that the high rooting rate ofP. crassifoliacuttings due to its ability to accumulate high concentrations of indole-3-acetic acid(IAA)and total carbon(TC)rather than abscisic acid(ABA)and total nitrogen(TN).The rooting rate was mainly regulated by the IAA/ABA and TC/TN ratio.In summary,our results suggest that the softwood cuttings(0.5–1.0 cm in diameter) from upper branches of 15 year-oldP.crassifoliacan be considered as an effective strategy to improve cutting rooting rate,and the IAA/ABA and TC/TN ratio was one of the main factors limiting the cutting rooting rate ofP. crassifolia.

Picea crassifoliaKom⋅Type⋅Position and age effect⋅Cutting reproduction⋅Rooting rate⋅Hormones⋅Organic substances

Introduction

Desertif i cation has been recognized as a major environmental problem that inevitably triggers a loss of soil productivity and a progressive reduction of vegetation coverage(Gomes et al.2003;Zhao et al.2009).The Qilian Mountains is one of the major mountains of northwest China,and its unique forest ecosystem plays a crucial role in the construction of the ecological environment(Guo et al.2003).However,the over and unreasonable use of forest resources has contributed to serious environmental degradation problems.Some plants,such asPicea crassifoliaKom andSabina przewalskii,have evolved various adaptive mechanisms that allow them to survive and grow well in those environments.In facing the challenge ofdesertif i cation,the physiological behaviors and adaptations involved in these species generate great interest(Zhang et al.2009;Liu et al.2012).

Picea crassifolia,a perennial arbor species,is recognized as one of the most adaptable plants found to date in Qilian Mountains(Duan 2012).This species is widely used to conserve soil and water in the barren mountain areas of northwest China.Meanwhile,the high productivity and excellent adaptability makeP.crassifoliaattractive as a garden ornamental species in local regions.At present, most researchers mainly concentrate on its growth and physiological characteristics,distribution patterns,and dynamics of population and sexual reproduction(Wang et al.2009;Zhao et al.2010;Meng et al.2011).

However,due to the alpine climate and low seed vigor, the sexual reproduction ofP.crassifoliais extremely restricted,leading to a short supply(Ji et al.1996;Chen et al.2007).Cutting reproduction has turned out to be a typical,convenient,and economical asexual reproduction technology,which has been widely used to cloneP. aspoerata(Ma 1993;Wu 2008).Results showed that the early growth rate ofP.aspoerataby cutting reproduction was signif i cantly higher than that of seedlings(Browne et al.1997;Bhardwa and Mlshra 1998),which cannot only keep the biological characteristics of ortets but also advance fruiting(Bengt 1981;Wang et al.2011).The Research Institute of Forestry Genetics,Columbia,British has begun to produce mass clones ofPicea sitchenrsis, which can resist the weevil parents(Talbert et al.1993). The cutting result ofP.abiesshowed that the rooting ability of different cutting types(length,diameter,position, age)was different(Bentzer 1988;Hannerz et al.1999),and the rooting ability gradually declined with the increase of forest age(Kleinschmit and Schmidt 1977).

In China,many studies have been conducted on related themes of cutting and vegetative propagation of conifers (conventional,dwarf colorful,new superior,ornamental species,etc.)(Du and Zhou 2007;Huang and Huang 2011; Du and Jia 2012;Fang et al.2013).The cutting results ofP. abiessuggest that the rooting numbers,total root length and the root-effect index of hardwood cuttings outnumbered softwood cuttings,and the cuttings length at＞15 cm is the best(Jin et al.2009).Wang(2008)analyzed the cutting propagation of four species ofPicea,indicating that the cutting rooting rate was positively correlated with cutting length,breeding substrates,agesofortetsand hormones,and with the following orderofPicealikiangensis＞Piceaglauca＞Picea mongolia＞P.crassifolia.Result also showed that softwood cuttings ofPieea koraiensistreated with different hormones and breeding substrates generated signif i cant rooting rates.The hormones effect following order of IBA＞NAA＞IAA＞ABT1＞CK and substrates effect order of forest soil＞peat soil＞perlite+peat soil(Liu et al.2011).

Little is known about the systematic cutting reproduction ofP.crassifolia,especially,the rooting mechanism.In this paper,we usedP.crassifoliato explore the optimal cutting reproduction technology,and also discuss and analyze the endogenous hormones and organic nutrients inP.crassifoliacuttings to reveal its physiological and biochemical basis.

Materials and methods

Site description

The study area(Fig.1)is located in Haxi Natural Protection Station(HNPS),Administration of Qilian Mountains Natural Nature Reserve,Gansu Province,Northwest China (37°16′–37°45′N,102°01′–102°51′E;elevation: 2100–4140 m).The variation in temperature amplitude of and uneven distribution of monthly precipitation are the major meteorological features in this region.The annual average temperature in the area is 1.2°C(extreme high temperature ranges from 28.5 to 32.4°C in July and low temperature varies from-27.8 to-29.0°C in January). On average,the annual mean sunshine hours is 1892 h, annual precipitation ranges from 401.9 to 632.3 mm,half of which comes in July and August and annual evaporation varies between 1041.2 and 1234.2 mm,and the relative humidity is about 58%(Wang and Che 2012).The mountain grey brownish soil(elevation:2400–3300 m) was the main distribution region of arbor forest(P.crassifolia,S.przewalskii).TheP.crassifoliais distributed in the shade and half-shade slopes,which account for 24.7% of the total area of water conservation forest and 75.7%of the total area of arbor forest in Qilian Mountains(Peng et al.2008).

Experimental design

Cutting type effect

Three cutting types(softwood:0.3–0.5 cm and 0.5–1.0 cm, hardwood:more than 1.0 cm and root:0.5–1.0 cm in diameter)and one cutting length(15 cm)were treated to select the most compatible cutting type ofP.crassifolia(April to October 2009).

The softwood cuttings were semi-lignif i ed branches with terminal buds,which were not affected by pests and diseases.Hardwood cuttings were strong branches with terminal buds,which were not affected by pests anddiseases.Root cuttings(main or lateral roots)were collected from healthy plants where the above-mentionedP. crassifolia(10 years-old)was naturally distributed.Furthermore,the polarity of the cuttings was considered.The upper part of polarity should be f l at,and the surface of cut should be 0.3–0.4 cm higher than the bud.The lower part should be a horseshoe-shaped oblique cut(in order to take root quickly)and the surface of cut should be 0.4–0.5 cm lower than the bud.To prevent cuttings from dehydrating by adverse circumstances,50 cuttings were bundled together and packed in a plastic bag to keep moist.

Fig.1 Location of Haxi Natural Protection Station (HNPS),Administration of Qilian Mountains Natural Nature Reserve,Gansu Province,northwest China (37°16′–37°45′N,102°01′–102°51′E)

The cutting experiment(f i eld culture)was conducted at a greenhouse in Haxi Natural Protection Station(HNPS) and the nursery site was subdivided into 16 similar plots under the same site conditions.Each plot(1 m length×1 m width)was surrounded by an isolated zone (0.2 m)and a seedbed was set up each plot(15 cm in height,1 m in width)in order to reduce water loss and improve survival rate of the cutting.After all preparatory work has been done(the lower oblique cuts of cuttings were soaked for 24 h in water),cuttings were removed the needles and cut in each plot(10 cm in plant space;15 cm in row space and 3–5 cm in cutting depth),and they were thoroughly watered.

The carbendazol(30–40 g/L)sprayed and resterilized on the seedbed once a week.Soil water content(SWC, 30 cm in depth)was maintained at 70%of f i eld water capacity(FWC)by weight;this value was maintained by irrigating with the 70%FWC during the experimental period.Each treatment consisted of four plots containing 50 plants each.After the plants were harvested,18 plants (survived,uniform plant)of each treatment were dug out and all plant tissues were washed three times with water for morphological analysis.

Cutting position and age effect

When the most compatible cutting type(softwood cuttings: 0.5–1.0 cm in diameter)was selected,we divided the whole tree into three equal parts(in addition to top branches)and de fi ned as upper,middle,and bottom branches from top to bottom.Four positions(top,upper,middle and bottom)and one cutting length(15 cm)of 10 year-old healthy plants were treated to select the most compatible cutting position ofP.crassifolia(May to October 2010). The detailed experimental process of the experiment was similar to that in different cutting positions ofP.crassifolia.After the most compatible cutting position(softwood: 0.5–1.0 cm in diameter)and type(upper branches)were found,the cuttings were collected from 7-,10-,15-,20-, and 25 year-old healthy ortets,which were treated to select the most compatible cutting age ofP.crassifolia(May to October 2011).The detailed experimental process of the experiment was similar to that in different cutting types and positions ofP.crassifolia.

Rooting mechanism of P.crassifolia cuttings

The experiment was conducted to determine whether the hormones or organic nutrients were one of the main factors limiting the cutting rooting rate ofP.crassifolia.We did this by discussing and analyzing the effect ofendogenous hormones(IAA,ABA and KT)and organic nutrients(TC and TN)(May to October 2012).One-year old softwood(0.5–1.0 cm in diameter)and hardwood (more than 1.0 cm in diameter)cuttings were collected from 7-,10-,15-,20-and 25 year-old healthy ortets where the above mentionedP.crassifoliawas naturally distributed.The phloem(20–30 g)of healthy softwood and hardwood cuttings was collected and placed in sealed plastic bags,and then taken to a laboratory for chemical analyses.

Measurements

Growth-related parameters

The rooting index of cuttings was determined,using routine methods after the growth stage ended.The mean root length(MRL)and total root length(TRL)were measured using a vernier caliper with 0.1 mm precision.The rooting rate(RR)was measured using the following formula: RR=the total numbers of rooted plants/the total numbers of cutted plants.

The root effect index(REI)was used to evaluate the rooting feature of various types of cuttings and the REI was estimated using the following formula:REI=(mean root length×the number of roots)/the total number of cuttings (Zhu and Wang 1991).

Endogenous hormones

The phloem(20–30 g)was collected according to the method described by Liang and Zhang(1997)and Buhtz et al.(2004).The endogenous hormones,including Indole-3-acetic acid(IAA);Abscisic Acid(ABA);and Kinetin(KT)were measured according to the method described by Li(2003).Brief l y,the IAA,ABA,and KT were measured by the method of high-performance liquid chromatography(HPLC).The chromatographic column includes:Novapak C18(0.4 cm×15 cm);mobile phase: 40%CH3OH-15%;CH3CN-45%H2O(volume fraction);pH 4(using H3PO4);speed:0.7 m/L min;and detector:UV254 nm×0.1AUFS.

Organic nutrients

The detailed collective method and process of phloem (20–30 g)were similar to the measurements of endogenous hormones.The organic nutrients,total carbon(TC) and nitrogen(TN),were measured by the method described in Guo et al.(2004).Brief l y,the concentration of TC was measured by the method of sodium thiosulfate titration and the TN was measured by Kjeldahl determination.

Statistical analysis

All data was analyzed following SPSS program for Windows Version 13.0(SPSS Inc.,Chicago,IL,USA,1975). Duncan’s multiple range tests and one-way analysis of variance(ANOVA)were used to compare the differences among the treatments.The least signif i cant difference (LSD)tests were performed to determine the signif i cance of treatment means atp＜0.05.

Results

Effect of different cutting types on rooting characteristics ofP.crassifolia(10 year-old)

The effect of different cutting types(softwood:0.3–0.5 cm and 0.5–1.0 cm,hardwood:more than 1.0 cm and root: 0.5–1.0 cm in diameter)on the cutting characteristics(REI, RR,MRN,MRL,TRL,and BRR)ofP.crassifoliahave reached a signif i cant level(p＜0.05),indicating that the different cutting types signif i cantly affected the cutting rooting ofP.crassifolia(Table 1).The rooting characteristics of softwood cuttings(0.5–1.0 cm in diameter) showed a better growth performance,and the RR was 28.1, 103.9,and 67.5%higher than that of softwood (0.3–0.5 cm in diameter),hardwood(more than 1.0 cm in diameter)and root(0.5–1.0 cm in diameter)respectively The MRN were 1.4–2.3 times than that of the softwood (0.3–0.5 cm in diameter)and hardwood(more than 1.0 cm in diameter)cuttings.The BRR were 22.9,37.3,and 74% lower than that of softwood(0.3–0.5 cm in diameter), hardwood and root cuttings,respectively(p＜0.05).

Effect of different cutting positions on rooting characteristics of softwood cuttings(0.5–1.0 cm in diameter)ofP.crassifolia(10 year-old)

The effects of different cutting positions(top,upper, middle and bottom)on the rooting characters(REI,RR, MRN,MRL,TRL and BRR)of softwood cuttings ofP. crassifoliahave reached signif i cant level(p＜0.05),indicating that the different cutting positions also signif i cantly affected the cutting rooting of softwood cuttingsof P. crassifolia(Table 2).The rooting rate of softwood cuttings from top and upper branches showed a better growth performance,and the RR were 32.1 and 19.5,49.8 and 37.2% higher than that of middle and bottom branches respectively.The MRN were 72 and 36,138.9,and 88.9%higher than that of middle and bottom branches,and especially, the BRR were 23.8 and 10.8,40.3 and 24.7%lower than that of middle and bottom branches respectively (p＜0.05).However,although the cutting rooting rate ofsoftwood cuttings from top branches was best,the tree structure was destroyed.

Table 1 The growth performance of P.crassifolia cuttings treated with different cutting types(softwood,hardwood and root)and one cutting lengths(15 cm)for 6 months(May to October 2009)

Table 2 The growth performance of P.crassifolia cuttings treated with different cutting positions and one cutting lengths(15 cm)for 6 months (May to October 2010)

Effect of different ortet ages on rooting characteristics of softwood cuttings(upper branches)ofP.crassifolia

The effects of different ortet ages(7-,10-,15-,20-,and 25 year-old)on the rooting characters(REI,RR,MRN, MRL,TRL,and BRR)of softwood cuttings(upper branches)ofP.crassifoliahave reached a signif i cant level (p＜0.05),indicating that the different ortet ages also signif i cantly affected the cutting rooting of softwood cuttings(upper branches)ofP.crassifolia(Table 3).The cutting rooting rate of softwood cuttings(upper branches) was increased f i rst and then decreased signif i cantly with the increasing age ofP.crassifolia.The cutting rooting rate of 15-year old ortets was better than that of 7-,10-,20-,and 25 year-old ortets(Table 3).The RR of 15 year-old ortets were 38.1,14.6,28.5,and 48.8%,higher than that of 7-, 10-,20-and 25-year-old ortets.The MRN were 82.9,41.5, 141.9,and 294.7%higher than that of 7-,10-,20-and25-year old ortets.Finally,the BRR were 36.9,12.7,26.2, and 47.4%lower than that of 7-,10-,20-,and 25 years-old ortets,respectively(p＜0.05).

Table 3 The growth performance of P.crassifolia cuttings treated with different ortet ages and one cutting lengths(15 cm)for 6 months(May to October 2011)

Table 4 Relationship between endogenous hormones and rooting rate of hardwood and softwood cuttings of P.crassifolia at different ages (May to October 2012)

Effect of endogenous hormones on rooting of hardwood and softwood cuttings of different ages ofP.crassifolia

The effect of endogenous hormones on the rooting of softwood and hardwood cuttings of different ages ofP. crassifolia(7-,10-,15-,20-,and 25 year-old)were obvious (Table 4).The rooting rate of hardwood and softwood cuttings of different ages ofP.crassifoliawere positively correlated with the ortet ages and IAA,ABA,and KT content.The content of IAA and KT in hardwood and softwood cuttings signif i cantly decreased,while the ABA was signif i cantly larger with the increasing age ofP. crassifolia.The content of IAA,ABA,and KT,and especially the ABA in hardwood cuttings,were signif i cantly higher than that in the softwood cuttings with the increasing age ofP.crassifolia.However,the IAA/ABA ratio has a positive effect(IAA/KT shows no obvious effect)on the rooting of hardwood and softwood cuttings with the increasing age ofP.crassifolia.The IAA/ABA ratio in softwood cuttings of 7-,10-,15-,20-and 25 yearoldP.crassifoliawas average 7.34 times more than that of the hardwood cuttings respectively(p＜0.05).In short,the rooting rate ofP.crassifoliacuttings was not decided by a single hormone but follows the IAA/ABA ratio.The bigger the ortets get,the smaller the IAA/ABA ratio has,and the lower rooting rate was either hardwood or softwood cuttings.

Effect of organic nutrients on rooting of hardwood and softwood cuttings of different ages ofP. crassifolia

The effect of organic nutrients on the rooting of softwood and hardwood cuttings of different ages ofP.crassifoliawere also obvious(Fig.2).The rooting rate of hardwood and softwood cuttings of different ages ofP.crassifoliawere positively correlated with the organic nutrient content,and the total carbon(TC)content in hardwood and softwood cuttings were decreased signif i cantly,while the total nitrogen(TN)was showed the opposite trend with the increasing age ofP.crassifolia(Fig.2).However,the C/N ratio has a positive effect on the rooting of hardwood and softwood cuttings with the increasing age ofP.crassifolia. The C/N ratio in softwood cuttings of 7-,10-,15-,20-and 25 year-oldP.crassifoliawas 97.4,119.5,106.5,82.7,and 103.8%higher than that in hardwood cuttings.In short,the rooting rate ofP.crassifoliacuttings was not decided by a single inclusion but by the C/N ratio.The bigger the ortets get,the smaller the C/N ratio becomes,and the lower the cuttings rate.

Discussion

The type,position,and age effect ofP.crassifoliaand its adaptation to alpine conditions

P.crassifoliais a perennial arbor species,and its growth condition is very complex resulting in its unique adaptation to alpine conditions.Our result showed that the amount of plant recruitment(picea)mainly relies on sexual reproduction to maintain its stability,and the growth performance of plants was extremely restricted due to their biological characteristics and habitat conditions which are far behind the asexual reproduction by cutting(Bengt 1981;Ma 1993).

In recent years,the sexual reproduction ofPiceaspecies is developing rapidly,and the application of various techniques in breeding(greenhouse seedling,container seedling,sand culture,etc.)shorten the breeding cycle ofPiceaspecies and improved the quality of seedlings(Cai 2010;Ji and Yao 2010;Kang et al.2010).However,the sexual reproduction ofP.crassifoliahas a slow growth rate fromthe seedling stage to the late fruit stage,a long time from fi rst building a garden to seed production,seed yield instability,and low economic eff i ciency(Zhang 1992;Ji et al.1996;Chen et al.2007;Lei et al.2012).Thus,the sexual reproduction ofP.crassifoliamay be virtually useless while the strong asexual reproductive performance plays an important role in mass production ofP.crassifolia(Dormling and Kellerstram 1981;Bentzer 1988).In this study,the cutting reproduction ofP.crassifoliahas type, position and age effect,and the softwood cuttings (0.5–1.0 cm in diameter)from the upper branches of a 15 year-old ortet showed a better growth performance on the cutting rooting ofP.crassifolia(Table 1,2 and 3).This is because the rooting rate of softwood cuttings (0.5–1.0 cm in diameter)from upper branches of 15 yearold ortet produces strong metabolism,high endogenous hormone contents,and vigorous meristematic ability of cells(Talbert et al.1993;Zhou 1995).

Fig.2 Relationship between total carbon(a),total nitrogen(b),and total carbon/total nitrogen ratio(c)and rooting rate of hardwood (H)and softwood(S)cuttings of P.crassifolia at different ages(May to October 2012)

It is noteworthy that the age effect on the cutting reproduction ofP.crassifoliahas an obvious aging effect, ref l ected in the decreased cutting rate,declined rooting quality,and slowed growth(Table 3),which is mainly due to the progress of individual development caused by the physiological changes of the growth point(Ma 1993). Similar results have been reported for some other plants, such asPicea koraiensis(Nakai)(Zhao et al.1997),Cunninghamia lanceolata(Lamb.)Hook(Shi et al.1993;Guo 1997),Pinus massoniana Lanb.(P.sinensis Lamb)(Ji et al. 1998)andPicea abiesKarsts(Talbert et al.1993;Hu et al. 2008;Jin et al.2009).From the countermeasures view of plants to adapt to environment,the asexual reproduction ofP.crassifoliais the dominant choice,having many advantages including short breeding cycle,fast growth, strong resistance,and stable genetics.Researchers have successfully reproduced younger,stronger,and healthier cuttings by reducing the height ofP.abiesfor asexual reproduction(Dormling and Kellerstram 1981;Clair et al. 1985).These results conf i rm that the evolutionary characteristics of asexual reproduction forP.crassifoliais an almost inevitable result under the environment stress.Each plant has its own complex physiological adaptive mechanisms that allow it to survive and grow well in a specif i c environment.

The effect of endogenesis hormones on the cutting rooting ofP.crassifolia

In 1934,Holland physiologist(Went F.W.)published the seminal paper that shows that hormones have positive effect on the adventitious root formation of plants,and since then,many scholars found that softwood cuttings with high endogenesis hormone content take roots easily. The winter dormancy branch,and the rooting rate of cuttings with bud and leaves is higher than the cuttings getting rid of bud and leaves(Guo 1997).This is because the endogenesis hormone—produced by buds and leaves through polar transport and accumulation in the base of cuttings—played on signif i cant role in rooting of cuttings.Chen et al.(2007)showed that endogenesis hormone, 6-benzyl aminopurine(6BA),on the remaining buds after apical buds were removed,was more effective in boosting the growth and development of buds and branches inP. crassifolia.

Duan et al.(2009)showed that plant growth regulators (ABT1)can signif i cantly promote the rooting of softwood cuttings with the order ofP.crassifolia＞P.asperata＞P. abies.Jin et al.(2009)showed that cuttings ofP.abiestreated with different hormones generated signif i cantly different rooting rates and with the effect following order of ABT1＞GGR＞IBA＞NAA.However,the number of roots generated,total rooting length,and the root effect index of the cuttings treated with IBA were better than those treated by other hormones.Moreover,being soaked two and half hourswith ABT1 at respective concentrations of 100 and 200 mg kg-1proved to be the best in terms of rooting rates,total rooting length,and the root effect index.

Liu et al.(2011)indicated that the rooting rates,growth condition of seedlings,and rooting period of softwood cuttings ofP.koraiensisobviously improved following treatment with 100 mg mL-1IBA,compared with those treated by other hormones and following the order of IBA＞NAA＞IAA＞ABT1＞CK.Similiar results have been reported onP.abiesand some otherPiceaspicies such asP.likiangensis,Picea glaucaandPicea koraiensiswhich showed that the rooting rate of cuttings was positively correlated with different hormone concentrations and immersion time(Hu et al.2008;Huang et al.2011;An et al.2011;Liu et al.2011;Hu et al.2014).

Our results showed that endogenous hormones play important roles in rooting ofP.crassifolia(Table 4):the IAA concentration and IAA/ABA ratio have positive correlation with cutting rooting rate,while the content of ABA has negative correlation with rooting rate(Table 4).This result shows that the low content of IAA(high content of ABA)made the IAA/ABA ratio very low,producing obviously inhibitory effects on the rooting of hardwood and softwood cuttings.These results are consistent with the results of Zheng(1991)and Zhang et al.(2004),which show that the IAA/ABA ratio plays an important role in cutting rooting of plants.Zhan et al.(2001)showed that the IAA and ABA contents play an important role in taking roots ofBetula platyphyllaSuk cuttings and the IAA/ABA ratio can be used as an important index to measure the cutting rooting ability.In short,the cutting rooting character ofP.crassifoliais not decided by a hormone or inclusions,but by the synergetic effect of endogenous hormone and the inclusion.The higher IAA content and lower ABA content made the IAA/ABA ratio very high, which may be one of the main reason leading to a higher cutting rooting rate in softwood cuttings and a lower cutting rooting rate in hardwood cuttings.

The effect of organic nutrients on the cutting rooting ofP.crassifolia

The C/N ratio ref l ects the nutritional utilization eff i ciency of plants and is an important supporter and regulator of plant-life processes.It is also one of the factors of the litter decomposition rate(Zhou et al.2004,2009).Healthy and developed cuttings are more likely to take root compared to cuttings with poor growth;clearly,the nutrient substances in cuttings may have a positive effect on the differentiation of the original body of roots(Guo 1997).Wang et al. (2011)analyzed the cutting propagation of three species ofPicea,indicating that the physiological age and nutrient content of cuttings had signif i cant correlation,and the nutrients,especially nitrogen(N)and potassium(K),in cuttings had great effects on the rooting of cuttings.

Our results showed that the cutting rooting rate ofP. crassifoliacorrelated to the C/N ratio,the high content of TC and low content of TN made the C/N ratio very high, which plays an important role in the cutting rate ofP. crassifolia,and the higher the C/N ratio is,the higher the cutting rooting rate gets.Moreover,the bigger the ortets get,the smaller the C/N ratio has,and the lower the cutting rooting rate was(Fig.2).Guo et al.(2004)showed that the total sugar(TS),total nitrogen(TN),the ratio of TS/TN was positively related to cutting rooting ofPinus bungeana.The results of Guo et al.(2004)are consistent with our results,which showed that the TS and ratio of TS/TN were closely correlated to cutting rooting ofPinus bungeana.Moreover,many scholars considered that the nutrient substances in cuttings are not the critical factors on the formation of adventitious roots of cuttings.Pinus bungeanais more likely to take roots when the proportional relationship among carbohydrates,nitrogenous compounds,and endogenous hormones reaches a certain level(Guo 1997).The rooting of cuttings may be largely dependent on the degree of branch development(collection time of cuttings).We can consult the morphological and anatomical characteristics(the elasticity,color,lignif i ed degree,and development degree of lenticels of branches, etc.)to carry out the cutting reproduction scientif i cally in a production setting.

Conclusions

Our results suggest that the softwood cuttings(0.5–1.0 cm in diameter)from upper branches of 15 year-oldP.crassifoliacan be considered as an effective strategy to promote cutting rooting rate and improve growth performance of roots.The IAA/ABA and TC/TN ratio inP.crassifoliacuttings was one of the main factors limiting the rooting rate ofP.crassifoliacuttings.We determined that threefactors—plant growth regulators,proper fertilization management of ortets,and bud-forcing treatment—can better improve the cutting rooting rate and growth performance ofP.crassifolia.They are innovative and open up a new perspective for the improvement ofP.crassifoliafor biomass production in Qilian Mountains.

AcknowledgmentsThe authors are grateful to the anonymous reviewers and editors for their critical review and comments that helped to clarify and improve the manuscript.

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28 October 2013/Accepted:18 August 2014/Published online:11 July 2015

Project funding:This work was supported by the National Natural Science Foundation of China(Grant No.31360086)and the grant from the major state basic research development program of china (973 program,No.2013CB429903).

The online version is available at http://www.springerlink.com

Corresponding editor:Hu Yanbo.

✉Wenzhi Zhao

zhaowzh@lzb.ac.cn

1Linze Inland River Basin Research Station,Key Laboratory of Inland River Basin Ecohydrology,Cold and Arid Regions Environmental and Engineering Research Institute,Chinese Academy of Science,Lanzhou 730000,Gansu,China

2Gansu Research Academy of Forestry Science and Technology,Lanzhou 730000,Gansu,China

3Haxi Natural Protection Station,Administration of Gansu Qilian Mountains National Nature Reserve, Wuwei 733000,Gansu,China