桃(Prunus persica(L.) Batsch.)品种核心种质的构建与评价

为构建桃品种核心种质,通过对56份桃（Prunus persica（L.） Batsch.）初级核心种质的形态农艺性状数据（MOR）和SSR等位基因数据的分析,研究了不同聚类取样方法和完全随机取样方法下9种取样比例的遗传多样性指数、保留比例及各频率段等位基因的丢失比例。结果表明：聚类取样的方法优于完全随机取样,并以在80%的取样比例下MOR结合SSR数据聚类取样的效果最好,利用此方案构建的桃品种核心种质共包括45份材料,该核心种质的基因遗传多样性指数最高,保留了初级核心种质100%的形态农艺性状和96.6%的SSR等位基因,在出现频率低于0.05的等位基因中共丢失了2个等位变异,保留了出现频率在0.05-0.10的所有等位基因;利用6个数量性状对所构建的核心种质的代表性检测表明所构建的核心种质很好地代表558份桃原始种质的遗传变异。

Molecular Identification and Cultivar Fingerprints of Prunus persica (L.) Batsch Germplasms

[Objective] The aim was to study the molecular identification and cultivar fingerprints of Prunus persica (L.) Batsch germplasms.[Method] Sixty peach genotypes,representing China common local cultivars and European samples were screened by microsatellites (simple sequence repeats,SSRs) and Inter-Simple Sequence Repeat (ISSR) markers.[Result] 26 reproducible bands were amplified by Nine SSR primers,and 24 of which were polymorphic; 236 bands were amplified by 30 ISSR primers,and 113 of which were polymorphic.31 genotypes were discriminated with 1-3 distinct polymorphic bands generated from the primers ISSR and SSR.Seven cultivar-specific ISSR fragments and two SSR unique alleles obtained from this study were available to be converted into Sequence Characterized Amplified Region (SCAR) markers.The genetic similarity coefficient (GS) estimated from these molecular data averaged were 0.939 (ranged from 0.856 to 0.983) for ISSR and 0.646 (ranged from 0.240 to 1.000) for SSR,respectively.The combined grouping association indicated that most local Chinese peach cultivars and exotic accessions were clustered together.This could be related to the mode of introduction and maintenance of the peach cultivars involving limited foundation germplasm,exchange of cultivars between plantations,and periodic development of new recombinant cultivars following sexual reproduction.[Conclusion] The results obtained in this work would help to improve the conservation,molecular identification and management of peach germplasm in breeding.

SSR方法标记桃[Prunus persic(L.)Batsch]果肉近核色素(英文)

[目的]利用 SSR 分子标记法标记桃(Prunus persic (L.) Batsch)果肉近核色素。[方法]以"重阳红"与"燕红"2 个桃品种为亲本构建正交 F1群体,选取其中138株后代作为标记群体,采用分离群体分组分析(bulked segregate analysis,BSA)法,将果肉近核色素分为"有"和"无"2个基因池,应用SSR分子标记技术寻找与桃果肉近核色素性状基因连锁的分子标记。[结果]通过对256对引物的筛选,获得了3对与控制桃果肉近核色素性状基因连锁的分子标记,即 UDP96-003、ch04g09 和 UDP97-402,同时计算得到这 3 个标记与桃果肉近核色素性状基因的遗传距离分别为16.7、10.1和17.0 cM。[结论]该研究为进一步筛选遗传距离更近的共显性分子标记奠定了基础。

Effects of nitrogen content on growth and hydraulic characteristics of peach(Prunus persica L.) seedlings under different soil moisture conditions

A pot experiment was conducted to investigate the effects of nitrogen content [Nl （no fertilizer）, N2 （0.15 g.kg-l）, and N3 （0.3 g.kg 1）] on the growth and the hydraulic characteristics of peach seedlings under different soil moisture conditions （Wl, W2 and W3, in which the soil water content was 45% to 55%, 60% to 70%, and 75% to 80% of the field water capacity, respectively） by using a specialized high pressure flow meter with a root chamber and a coupling, which was connected to plant organs. Leaf area and leaf hydraulic conductivity （KL） increased significantly in the seedlings because of increased soil moisture and N content. KL increased with leaf area. A linear correlation was documented between KL and leaf area. KL was higher in the morning and began to decline sharply after 16：00, at which KL declined after an initial increase. Soil moisture and N content enhanced shoot （Ks） and root （Kr） hydraulic conductivities, thereby improving the low soil moisture condition to a large extent. Ks and Kr of the seedlings were reduced by 32% and 27% respectively in N~, and by 14.7% and 9.4%, respectively in N2, and both in Wb compared with the control treatment. N3 had no significant effect on Ks and Kr under similar conditions. Linear negative correlations were observed between Kr and the excised root diameter as well as between Ks and the shoot stem diameter. The shoot-to-root ratio increased with in- crease in N content. The shoot-to-root ratio in N3 was increased by 14.37%, compared with N1 in W1 as well as by 12% and 4.39% in Wz and W3, respectively. Knowledge of the effects of soil moisture and N fertilizer on hydraulic characteristics and growth is important. Our results provide basic guidelines for the implementation of water-saving irrigation and fertilization management of nursery stock.

Novel in silico EST-SSR markers and bioinformatic approaches to detect genetic variation among peach(Prunus persica L.)germplasm

Because there are thousands of peach cultivars,cultivar classification is a critical step before starting a breeding project.Various molecular markers such as simple sequence repeats(SSRs)can be used.In this study,67 polymorphic primers produced 302 bands.Higher values for SI index(1.903)suggested higher genetic variability in the genotype under investigation.Mean values for observed alleles(Na),expected heterozygosity(He),effective alleles(Ne),Nei’s information index(h),and polymorphic information content(PIC)were 4.5,0.83,5.45,0.83,and 0.81,respectively.The dendrogram constructed based on Jaccard’s similarity coefficients outlined four distinct clusters in the entire germplasm.In addition,an analysis of molecular variance(AMOVA)showed that70.68%of the total variation was due to within-population variation,while 29.32%was due to variation among populations.According to this research,all primers were successfully used for the peach accessions.The EST-SSR markers should be useful in peach breeding programs and other research.

桃(Prunus persica [L.] Batsch)转录组SSR信息分析及其分子标记开发

利用MISA软件筛选桃(Prunus persica[L.]Batsch)转录组数据,获得的109 248条Unigene,检测出24 102个SSR位点,分布于22 689条Unigene中,出现频率为20.76%。SSRs位点中主导类型是以AG/CT(占总SSRs的18.70%)为主的二核苷酸重复,占总SSRs的33%;其次是三核苷酸重复,出现频率为27%;四核苷酸重复,占总SSRs的25%。利用Primer 5共设计出9 657对SSR引物。随机选择50对引物进行PCR扩增,其中20对扩增出清晰可重复的条带,并且在同属材料(苹果试材)中也能获得理想结果,引物多态性检测发现,在15份桃试材中,10对引物具有多态性,利用UPGMA作图,可将15个桃材料很好区分。结果表明,桃转录组测序产生的Unigene是开发SSR标记的有效信息,为SSR标记在桃的遗传分析上提供可靠的标记选择。

Identification and expression analysis of chlorogenic acid biosynthesis key gene PpHCT in peach

Shikimic acid/quinic acid hydroxy cinnamyl transferase(HCT)is one of the key enzymes in the phenylpropanoid pathway.However,the role of the HCT gene in chlorogenic acid(CGA)biosynthesis in peach fruit remains unclear.For this,we identified the accumulation pattern of CGA in four peach cultivars,cloned and characterized 11 PpHCT gene members,and further analyzed the expression patterns of these PpHCT genes during fruit development.The contents of CGAs in the four peach cultivars all exhibited a trend of increasing and then decreasing during the fruit growth and development.Moreover,the contents of CGAs in the peel and flesh were tissue-specific.Gene structure analysis indicated that the PpHCT genes were highly conserved,containing two exons and one intron.The protein structure analysis demonstrated that the PpHCT proteins contained two conserved motifs(HXXXD,DFGWG)and a transferase domain(PF02458),which belonged to the BAHD acyltransferase family.The cis-acting element analysis suggested that the promoters of PpHCT genes contained many light-related,hormone-related,stress-related,tissue-specific,and circadian-related elements,and they could participate in a variety of biological processes.Phylogenetic analysis showed that the HCT proteins of peach were closely related to the HCT proteins of plum and had a close evolutionary relationship.The qRT-PCR analysis indicated that the expression levels of PpHCT1 and PpHCT2 showed an opposite trend to the accumulation of CGA,whereas the expression levels of PpHCT4,PpHCT5,PpHCT7,PpHCT8,and PpHCT11 demonstrated the same trend as CGA accumulation.It was worth noting that only PpHCT4 and PpHCT5 were highly expressed in the two high-CGA cultivars but showed low levels of expression in the two low-CGA cultivars.Therefore,it was hypothesized that these two genes might be key genes to the synthesis of CGA in peach fruit.Those findings provide a theoretical basis for further study on the biological functions of the HCT gene and help to reveal the molecular mechanism of CGA.

HPLC法测定不同产地桃仁中苦杏仁苷的含量

目的:建立不同产地桃仁中苦杏仁苷含量的高效液相测定方法,测定不同产地桃仁药材中苦杏仁苷的含量。方法:色谱柱HypersilBDS C18(250mm×4.6mm,5μm),流动相为乙腈-水(17∶83)。结果:苦杏仁苷在4.94～80μg/mL范围内线性关系良好,r=0.9996,平均回收率为99.87%,RSD=1.01%。含量测定结果表明,不同产地桃仁药材中苦杏仁苷含量存在差异。结论:该方法简便快捷,准确灵敏,重复性好,可用于桃仁药材的质量控制。

不同方法测定桃树抗寒性关系研究

以生产中抗寒性表现较强、中等、较弱的3个桃品种为试材,应用电解质渗透法(EL)和电阻抗图谱法(EIS)研究了不同时期枝条抗寒性和未经冷冻处理枝条EIS参数的变化,并对未经冷冻处理枝条EIS参数、冷冻处理枝条EIS参数测定的半致死温度与EL测定的半致死温度的关系进行了统计分析。结果表明:未经冷冻处理3个桃品种的胞外电阻率(r_e)和胞内电阻率(r_i)均随着外界温度的降低而升高,随着外界温度的升高而降低;弛豫时间(τ)与r_(e)、r_(i)变化趋势相反;通过对4个电阻抗参数与EL测定的半致死温度统计分析得出,EIS(r_(i))是最佳参数,其相关系数为-0.828,通径系数为-7.385,线性回归方程为y=-4.063 9x+4.333 8,R^(2)=0.686。EL和EIS参数法测定3个桃品种的抗寒性与生产上抗寒性强弱表现一致;通过对2种方法统计分析得出,EL与EIS 4个参数法测定抗寒性均呈极显著正相关,其中弛豫时间法相关系数、通径系数和决定系数均最高,因此,弛豫时间是预测冷冻处理桃树枝条抗寒性的适宜参数。

Relationships Between the Distribution of Relative Canopy Light Intensity and the Peach Yield and Quality

The aim of the present experiment was to study the relationship between the distribution of relative light intensity in canopy and yield and quality of Wanmi peach. The optimum relative canopy light intensity was judged to be 36.3% for high quality peaches, when canopy volumes of Wanmi peach trees with a relative light intensity 〈 30% accounted for 7.7 and 47.9% of the total canopy volume in June and September, respectively. The canopy volume with a relative light intensity 〉 80% was 27.7 and 3.1% of the total canopy volume in June and September, respectively. Peach canopies were divided into 0.5 m × 0.5 m × 0.5 m cubes, with the relative light intensity being measured at different positions of the canopy during the growing season. Yield and fruit quality were also measured at these positions at harvest. The results showed that the relative light intensity decreased gradually from outside to inside and from top to bottom of the tree canopy. Fruit were mainly distributed in the upper and middle portions of the canopy, 1.5-3.0 m above ground. Regression results showed that single fruit weight and soluble solid content were positively related to relative light intensity.

Effects of Temperature and Several Chemicals on Metabolic Changes During Dormancy Release in NJ72 Nectarine

Poor, delayed and ununiform budbreak is a major problem for peaches in greenhouse. To clarify the mechanism of breaking bud dormancy in nectarines, the effect of temperature and three dormancy-breaking agents on metabolic changes during dormancy release in two-year old NJ72 nectarine (Prunus persica L. Batch) was investigated. The result showed temperature and chemicals affected the budbreak and the metabolism of NJ72 nectarine during dormancy. Endogeneous peroxide content in buds increased soon after low temperature treatment. Meanwhile, catalase activity was also shown to increase significantly at low temperature treatment, coincided with increase of the activity of peroxidase and superoxide dismutase. The rate of respiration in flower buds increased at low temperature during dormancy. The rate of the pentose phosphate pathway increased, while the rate of the Embden-Meyerhof pathway decreased and the rate of tricarboxlic acid cycle changed little. Glucose 6-phosphate dehydrogenase activity increased at low temperature during dormancy. At the same time we found an accumulation of peroxide after treatment with dormancy-breaking chemicals. In flower buds treated with dormancy-breaking agents, thiourea, KNO3 and NH4NO3, catalase activity was inhibited soon after treatment, whereas peroxidase activity increased, and the changes of superoxide dismutase remained little. In this study, it was found that the rates of respiration in flower buds increased by chemicals sprays during dormancy. The activity of glucose 6-phosphate dehydrogenase, the key enzyme in the pentose phosphate pathway (PPP), increased by spraying with dormancy-breaking agents, concomitantly with the activation of the pentose phosphate pathway.

Effect and Functional Mechanism of the Action of Exogenous Gibberellin on Flowering of Peach

This study was conducted to assess the effect of gibberellin and its possible mechanism of action on peach flower formation. At flower induction, 100 mg L^-1 of gibberellic acid 3 （GA3） was sprayed on the leaves of peach [Prunus persica （L.） Batsch.] cv. Bayuecui. Using anatomy, immunohistochemistry, and semi-quantitation, the in situ distribution of GAs and the expression of the key genes involved in peach flower formation in the apical meristem were studied during flowering differentiation. The results showed that induction of flowering in the Bayuecui peach occurred prior to 10 July in Beijing, China. Flower induction and further differentiation of the peach flower organs were significantly inhibited by leaf-spraying of GA3 at a concentration of 100 mg L^-1 during the induction stage. The flowering rate was only 11.67% after treatment. The distribution of GA1 in the apical meristem varied during the process of flower bud differentiation. From 13 June to 25 July, the GA1 signal from control plants was detected mainly in the vascular bundles at the base of the flower buds. No GA1 signal was detected in the apical meristem. After treatment with GA3, the distribution was similar to that of the control from 13 June to 3 July. On 13 July, a GA1 signal was detected in the apical meristem accompanied by an increase in the GA1 signal in the vascular bundles at the base of the flower buds. The GA1 signal weakened significantly in both the vascular bundles and the apical meristem on 25 July. The expression of the genes PpLEAFY and MADS6 in flower buds could be detected only on 10 October in the GA3-treated plants. The critical period for flower induction of Bayuecui peach in Beijing was in early July, during which time, leaf-spraying with 100 mg L-1 GA3 could effectively inhibit flower induction and further differentiation of the flower buds. GA1 in the gibberellin family was the suppressor for flower induction in peach. Its action was affected by the stage of flower bud differentiation. Expression of the key genes PpLEAFY and MADS6 involved in flower formation was inhibited by GA3 treatment.

氨基酸水溶肥促进桃苗生长

氨基酸水溶肥是具有营养效果好、无污染等优点的新型绿色肥料。以桃(Prunus persica L.)苗为材料,通过盆栽试验探究施用不同稀释倍数(600、900、1200、1500倍)的氨基酸水溶肥对桃苗生长的影响,以期筛选出效果最佳的喷施浓度。结果表明,施用氨基酸水溶肥增加了桃苗各部位的生物量,其根、茎、叶及地上部分生物量在稀释倍数为1500倍时最高,较对照分别增加了54.83%、51.28%、35.73%和42.12%;施用氨基酸水溶肥提高了桃苗光合色素含量,当稀释倍数为1500倍时,桃苗的光合色素含量最大;施用氨基酸水溶肥提高了桃苗的抗氧化酶活性,但降低了桃苗的可溶性蛋白质含量。因此,稀释1500倍的氨基酸水溶肥为促进桃苗生长的最佳喷施浓度。

丛枝菌根真菌对桃幼苗生长及硒富集的影响

采用盆栽试验,研究了摩西球囊霉(Glomus mosseae)、脆无梗囊霉(Acaulospora delicata)、隐类球囊霉(Paraglomus occultum)和幼套球囊霉(Glomus etunicatum)4种丛枝菌根真菌对桃[Prunus persica(L.)Batsch]幼苗生长及硒富集特性的影响。结果表明,施用脆无梗囊霉增加了桃幼苗的生物量,促进了桃幼苗的生长,使桃幼苗的根系和地上部分生物量分别比未施用增加了11.28%和9.18%,也在一定程度上提高了光合色素含量、过氧化酶活性及过氧化氢酶活性。施用摩西球囊霉、隐类球囊霉和幼套球囊霉3种丛枝菌根真菌降低了桃幼苗的生物量、光合色素含量及抗氧化酶活性或对其没有显著影响。施用脆无梗囊霉也在一定程度上增加了桃幼苗的总硒含量及有机硒含量,其中地上部分的总硒含量和有机硒含量分别较未施用增加了8.01%和9.34%。施用摩西球嚢霉、隐类球囊霉和幼套球囊霉降低了桃幼苗的总硒含量及有机硒含量或对其没有显著影响。

中国桃主要品种资源及其野生近缘种的分子身份证构建

【目的】以国家果树种质郑州葡萄、桃圃保存的237份中国桃地方品种、育成品种及其野生近缘种为试材,进行种质分子身份证构建工作,并对构建方法进行探讨。【方法】采用筛选后的SSR标记对品种进行区分,然后根据引物对不同品种扩增条带分子量的大小进行编码、组合。【结果】从80对引物中筛选出来自桃8条染色体上的16对SSR引物,在供试种质间共检测出等位基因203个,每对引物平均检测到等位基因数为12.7个。根据等位基因既位于地方品种、育成品种,又位于近缘野生种的选择方法,筛选出123对等位基因并赋值后可用于构建种质的分子身份证。【结论】在237份种质中有202份具有的可辨分子身份证编码,继续对不同引物组合对种质的筛选效率进行分析,筛选出8对核心引物使176份种质具有可辨的分子身份证,平均每对引物区分种质达22.1份,即在保证每对引物区分效率较高的基础上,兼顾了总的区分数量。最后,为使区分品种简便化,依据引物的多样性指数先后选择引物组合,达到用最少的引物组合区分不同品种的目的。

桃炭疽病病原鉴定及木醋液防治研究

炭疽病是湖北省桃[Prunus persica(L.)Batsch]产区一种重要的桃树病害。为有效控制该病的危害,试验采用分子生物学技术对湖北省桃园炭疽病的病原进行了分子鉴定,并在实验室内测定了不同浓度的木醋液对炭疽病病原菌的抑菌效果。结果表明,木醋液稀释300倍后对炭疽病病原菌的抑菌效果仍在70%以上;同时田间试验结果表明,木醋液能较好地控制炭疽病危害,在桃园绿色防控上具有广阔的应用前景。

湖北地方红肉桃种质资源收集、评价及利用

红肉桃[Prunus persica(L.)Batsch.]是湖北省的地方特色桃品种,其果实成熟时果肉呈紫红色或鲜红色,风味浓甜,质地脆爽,离核,在桃果肉颜色的更新选育上具有重要作用。从湖北红肉桃的特征特性、资源收集及评价、遗传育种、果肉颜色、红色素开发利用等几个方面进行了综述,并结合作者的研究工作情况对红肉桃育种利用及遗传倾向进行了讨论,分析了湖北红肉桃育种利用中存在的问题及难点。

桃分子连锁图的构建与分析

以‘大久保’与‘兴津油桃’杂交的F2 代 10 9株群体为试材 ,采用AFLP、RAPD、SSR分子标记进行遗传分析。筛选扩增稳定、多态性丰富的 36对AFLP引物、 3对SSR引物、 2对RAPD引物进行群体分离分析 ,获得分离标记 136个 ,卡方检验 2 7个标记偏离孟德尔分离比例。应用Mapmaker分析软件将符合孟德尔遗传分离比例的标记构建了包含 11个连锁群的连锁图谱 ,每个连锁群包含 3～ 2 1个标记 ,平均为 8 73个标记。该图谱覆盖基因组 10 6 1 8cM ,11个连锁群的平均长度为 96 5cM ,标记间平均图距为11 0cM ,与果实毛 /油桃 (G/g)、白 /黄肉 (Y/y)连锁的RAPD标记、非酸 /酸 (D/d)性状连锁的AFLP标记分别定位在第 3、 7、 9连锁群上。

桃园秋冬季生草栽培适宜草种的筛选与评价

南方果园大多集中在丘陵山区地带,土壤较为瘠薄,有机质含量较为低下。试验选择大麦草（Hordeum vulgare L.）、黑麦草（Lolium perenne L.）、毛叶苕子（Vicia villosa Roth.）等3个草种8个品种为研究对象,在桃[Prunus persica（L.）Batsch]园开展间作试验,对几个草种的耐旱性、耐寒性、封行期、观青期等特性进行了调查,测定了高度、盖度、重量等生物学指标。结果表明,冷季型草种大麦草综合性状较好,其抗寒性、抗旱性较强,第二年春季温度回升后生长快,行间封行早,刈割后再生能力较强,耐土壤贫瘠能力最强;并且大麦草生长期与桃园的冬闲期重叠时间最长,可达2~3个月,观青期长,可较好地改变桃园冬季落叶后的季相。因此,确定大麦草作为桃园秋冬季间作的适宜草种,这为湖北省乃至南方地区的桃园生草栽培技术应用提供了依据。

外源赤霉素对桃的成花效应及其作用机制

【目的】揭示赤霉素对桃成花的影响及其可能的作用机制。【方法】在桃花诱导期叶面喷施100mg·L-1GA3,运用解剖学、免疫组织化学及半定量RT-PCR方法,研究‘八月脆’桃花芽分化过程中顶端分生组织赤霉素的原位分布及成花关键基因表达变化。【结果】‘八月脆’桃在7月10日前处于成花诱导期(北京);成花诱导期叶面喷施100mg·L-1GA3显著抑制了桃花诱导及进一步的正常分化,处理后成花率仅为11.67%;顶端分生组织中GA1的分布随花芽分化进程而变化,6月13日-7月25日,花芽中GA1主要分布在芽基部的维管组织中,顶端分生组织中没有检测到信号。GA3处理后,GA1的分布在6月13日-7月3日与正常花芽相同,在7月13日顶端分生组织中检测到了GA1信号,且芽基部维管组织中的GA1信号增强,但至7月25日顶端分生组织和芽基部维管组织中的GA1信号明显减弱;GA3处理后,芽中PpLEAFY及MADS6基因仅在10月10日低量表达,其它时期几乎检测不到。【结论】在北京,‘八月脆’桃成花诱导期在7月10日之前;成花诱导期100mg·L-1GA3处理能显著抑制花诱导和进一步正常分化;赤霉素家族中GA1类在桃成花过程中起抑制作用,并且具有一定的时期性;GA3处理抑制了成花关键基因PpLEAFY及MADS6的正常表达。