The red bayberry genome and genetic basis of sex determination.

Morella rubra, red bayberry, is an economically important fruit tree in south China. Here, we assembled the first high-quality genome for both a female and a male individual of red bayberry. The genome size was 313-Mb, and 90% sequences were assembled into eight pseudo chromosome molecules, with 32 493 predicted genes. By whole-genome comparison between the female and male and association analysis with sequences of bulked and individual DNA samples from female and male, a 59-Kb region determining female was identified and located on distal end of pseudochromosome 8, which contains abundant transposable element and seven putative genes, four of them are related to sex floral development. This 59-Kb female-specific region was likely to be derived from duplication and rearrangement of paralogous genes and retained non-recombinant in the female-specific region. Sex-specific molecular markers developed from candidate genes co-segregated with sex in a genetically diverse female and male germplasm. We propose sex determination follow the ZW model of female heterogamety. The genome sequence of red bayberry provides a valuable resource for plant sex chromosome evolution and also provides important insights for molecular biology, genetics and modern breeding in Myricaceae family.

[Subtypes of muscarinic acetylcholine receptors involved in the persistent activity of layer V pyramidal neurons in the primary auditory cortex of young mice].

Cholinergic receptor activation and intracellular current injection lead to the persistent activity (PA), which may be involved in inducing neural plasticity. Our previous study showed that PA is closely related to the activation of muscarinic acetylcholine receptors (mAChRs) in pyramidal neurons of mouse primary auditory cortex (AI). However, the subtypes of mAChRs involved in PA remain unclear. Thus, using whole-cell patch-clamp recording and pharmacological methods, we investigated the role of different mAChR subtypes in inducing PA in AI layer V pyramidal neurons of young mice. The results showed that activation of mAChRs with intracellular depolarizing current induced PA in layer V pyramidal neurons. Blockade of M1, M2 or M3 subtypes prevented the PA, whereas M4 receptor antagonists did not affect the production of PA. The results suggest that the PA may be induced through a mechanism involving M1, M2 and M3 muscarinic receptors, but not M4 subtype.

Genetic diversity of male and female Chinese bayberry (Myrica rubra) populations and identification of sex-associated markers.

BACKGROUND: Chinese bayberry (Myrica rubra Sieb. & Zucc.) is an important subtropical evergreen fruit tree in southern China. Generally dioecious, the female plants are cultivated for fruit and have been studied extensively, but male plants have received very little attention. Knowledge of males may have a major impact on conservation and genetic improvement as well as on breeding. Using 84 polymorphic SSRs, we genotyped 213 M. rubra individuals (99 male individuals, 113 female varieties and 1 monoecious) and compared the difference in genetic diversity between the female and the male populations. RESULTS: Neighbour-joining cluster analysis separated M. rubra from three related species, and the male from female populations within M. rubra. By structure analysis, 178 M. rubra accessions were assigned to two subpopulations: Male dominated (98) and Female dominated (80). The well-known cultivars 'Biqi' and 'Dongkui', and the landraces 'Fenhong' are derived from three different gene pools. Female population had a slightly higher values of genetic diversity parameters (such as number of alleles and heterozygosity) than the male population, but not significantly different. The SSR loci ZJU062 and ZJU130 showed an empirical Fst value of 0.455 and 0.333, respectively, which are significantly above the 95 % confidence level, indicating that they are outlier loci related to sex separation. CONCLUSION: The male and female populations of Chinese bayberry have similar genetic diversity in terms of average number of alleles and level of heterozygosity, but were clearly separated by genetic structure analysis due to two markers associated with sex type, ZJU062 and ZJU130. Zhejiang Province China could be the centre of diversity of M. rubra in China, with wide genetic diversity coverage; and the two representative cultivars 'Biqi' and 'Dongkui', and one landrace 'Fenhong' in three female subpopulations. This research provides genetic information on male and female Chinese bayberry and will act as a reference for breeding programs.

[In vitro whole-cell patch clamp recordings of neurons in subnuclei of mouse inferior colliculus].

The inferior colliculus (IC) is a pivot along the central auditory pathway. Using infrared visual whole-cell patch clamp recording technique, we investigated the electrophysiological properties of IC subnuclei neurons. Recordings were made from 88 neurons, including 21 neurons from the dorsal cortex of the IC (ICd), 43 neurons from the central nucleus of the IC (ICc) and 24 neurons from the external cortex of the IC (ICx). Based on the responses to positive current injection, three firing patterns, i.e., onset (6.8%, n = 6), adapting (39.8%, n = 35) and sustained (53.4%, n = 47) patterns, were identified. The hyperpolarization-activated inward current (Ih) could be recorded in half of the neurons (49/88). The sustained pattern occurred in more than half of ICd and ICc neurons (61.9% and 67.4%), while the adapting pattern occurred in majority of ICx neurons (75%). Action potential (AP) threshold and time constant also showed significant differences across neurons from the ICd, the ICc and the ICx. Our results indicate that IC neurons are different in electrophysiological properties across the subnuclei. The variance of the responses may be related to the distinct types of neurons as well as the received projections, which is implicated in the distinct roles of IC neurons in central auditory processing.

Peach genetic resources: diversity, population structure and linkage disequilibrium.

BACKGROUND: Peach (Prunus persica (L.) Batsch) is one of the most important model fruits in the Rosaceae family. Native to the west of China, where peach has been domesticated for more than 4,000 years, its cultivation spread from China to Persia, Mediterranean countries and to America. Chinese peach has had a major impact on international peach breeding programs due to its high genetic diversity. In this research, we used 48 highly polymorphic SSRs, distributed over the peach genome, to investigate the difference in genetic diversity, and linkage disequilibrium (LD) among Chinese cultivars, and North American and European cultivars, and the evolution of current peach cultivars. RESULTS: In total, 588 alleles were obtained with 48 SSRs on 653 peach accessions, giving an average of 12.25 alleles per locus. In general, the average value of observed heterozygosity (0.47) was lower than the expected heterozygosity (0.60). The separate analysis of groups of accessions according to their origin or reproductive strategies showed greater variability in Oriental cultivars, mainly due to the high level of heterozygosity in Chinese landraces. Genetic distance analysis clustered the cultivars into two main groups: one included four wild related Prunus, and the other included most of the Oriental and Occidental landraces and breeding cultivars. STRUCTURE analysis assigned 469 accessions to three subpopulations: Oriental (234), Occidental (174), and Landraces (61). Nested STRUCTURE analysis divided the Oriental subpopulation into two different subpopulations: 'Yu Lu' and 'Hakuho'. The Occidental breeding subpopulation was also subdivided into nectarine and peach subpopulations. Linkage disequilibrium (LD) analysis in each of these subpopulations showed that the percentage of linked (r2 > 0.1) intra-chromosome comparisons ranged between 14% and 47%. LD decayed faster in Oriental (1,196 Kbp) than in Occidental (2,687 Kbp) samples. In the 'Yu Lu' subpopulation there was considerable LD extension while no variation of LD with physical distance was observed in the landraces. From the first STRUCTURE result, LG1 had the greatest proportion of alleles in LD within all three subpopulations. CONCLUSIONS: Our study demonstrates a high level of genetic diversity and relatively fast decay of LD in the Oriental peach breeding program. Inclusion of Chinese landraces will have a greater effect on increasing genetic diversity in Occidental breeding programs. Fingerprinting with genotype data for all 658 cultivars will be used for accession management in different germplasms. A higher density of markers are needed for association mapping in Oriental germplasm due to the low extension of LD. Population structure and evaluation of LD provides valuable information for GWAS experiment design in peach.

[Peach genomics and genome-wide association study: a review].

Peach (Prunus persica (L.) Batsch) is one of the most predominant stone fruits in Rosaceae family. The broad climate adaption, diverse cultivation region and good fruit taste make it one of the favorate fruits by consumers. Improving fruit quality and enhancing disease/pest resistance are always a focus for peach genetists and breeders to follow with interests. This paper reviews the main achievements on linkage map and physical map construction, development of various molecular markers, whole genome sequencing and transcriptome sequencing for peach in recent years, and also elaborates the applications of genome wide association study (GWAS) with high density SNP markers in peach and other plant crops. This review also provides a theoretical basis for GWAS analysis in the future study to identify high efficient markers of targeted traits for peach.

Development of simple sequence repeat (SSR) markers from a genome survey of Chinese bayberry (Myrica rubra).

BACKGROUND: Chinese bayberry (Myrica rubra Sieb. and Zucc.) is a subtropical evergreen tree originating in China. It has been cultivated in southern China for several thousand years, and annual production has reached 1.1 million tons. The taste and high level of health promoting characters identified in the fruit in recent years has stimulated its extension in China and introduction to Australia. A limited number of co-dominant markers have been developed and applied in genetic diversity and identity studies. Here we report, for the first time, a survey of whole genome shotgun data to develop a large number of simple sequence repeat (SSR) markers to analyse the genetic diversity of the common cultivated Chinese bayberry and the relationship with three other Myrica species. RESULTS: The whole genome shotgun survey of Chinese bayberry produced 9.01Gb of sequence data, about 26x coverage of the estimated genome size of 323 Mb. The genome sequences were highly heterozygous, but with little duplication. From the initial assembled scaffold covering 255 Mb sequence data, 28,602 SSRs (≥5 repeats) were identified. Dinucleotide was the most common repeat motif with a frequency of 84.73%, followed by 13.78% trinucleotide, 1.34% tetranucleotide, 0.12% pentanucleotide and 0.04% hexanucleotide. From 600 primer pairs, 186 polymorphic SSRs were developed. Of these, 158 were used to screen 29 Chinese bayberry accessions and three other Myrica species: 91.14%, 89.87% and 46.84% SSRs could be used in Myrica adenophora, Myrica nana and Myrica cerifera, respectively. The UPGMA dendrogram tree showed that cultivated Myrica rubra is closely related to Myrica adenophora and Myrica nana, originating in southwest China, and very distantly related to Myrica cerifera, originating in America. These markers can be used in the construction of a linkage map and for genetic diversity studies in Myrica species. CONCLUSION: Myrica rubra has a small genome of about 323 Mb with a high level of heterozygosity. A large number of SSRs were identified, and 158 polymorphic SSR markers developed, 91% of which can be transferred to other Myrica species.

[Effects of Bacillus fertilizer and agent on growth of young peach tree and soil environment under replant condition.] / èŠ½å­¢æ†èŒèŒè‚¥å’ŒèŒå‰‚å¯¹è¿žä½œæ¡ä»¶ä¸‹æ¡ƒå¹¼æ ‘ç”Ÿé•¿å’ŒåœŸå£¤çŽ¯å¢ƒçš„å½±å“.

We analyzed the effects of Bacillus fertilizer and agent supplementation at different concentrations in the replanted soil by examining the growth of young peach trees, soil nutrients, enzyme activities, and fungal commu-nity. The aim of this study was to provide theoretical basis for alleviating peach replant diseases. One-year-old potted 'Yuanmeng' peach trees were subjected to eight treatments: replanted soil without supplementation (RS), non-replanted soil without supplementation (NS), replanted soil with 1%, 4%, 8% Bacillus fertilizer supplementation (BF1, BF2, BF3), and 0.1‰, 0.5‰, 1‰ Bacillus agent supplementation (B1, B2, B3), respectively. The results showed that shoot growth and root biomass under NS treatment were greater than that under other treatments, and the contents of soil organic matter, available nutrients and soil catalase activity were greatly enhanced. Peach trees treated with BF and B treatments showed better root development than RS, among which, BF3 and B1 treatments were the best, and B1 treatment had similar effects as NS treatment. Compared with RS, BF and B treatments significantly increased the activities of soil catalase and urease during new shoot development, while B treatment significantly increased soil sucrase activity after the growth cessation of new shoots. Bacillus fertilizer and agent increased soil fungal diversity and the relative abundance of Ascomycota, Chaetomium and Penicillium, and decreased that of Basidiomycota. Considering the comprehensive parameters, BF3 and B1 showed more significant effects. In summary, our findings indicated that the supplementation of Bacillus fertilizer and agent to replanted soil could improve soil fertility, fungal community, and root development of roots, and thus provided a feasible method to alleviate peach replant diseases.

Construction of an anchoring SSR marker genetic linkage map and detection of a sex-linked region in two dioecious populations of red bayberry.

Red bayberry (Morella rubra) is an evergreen fruit tree found in southern China whose whole-genome sequence has recently been published. We updated the linkage map of the species by adding 118 SSR markers and the female-specific marker MrFT2_BD-SEX. The integrated map included eight linkage groups and spanned 491 cM. Eleven sex-associated markers were identified, six of which were located in linkage group 8, in agreement with the previously reported location of the sex-determining region. The MrFT2_BD-SEX marker was genotyped in 203 cultivated accessions. Among the females of the accessions, we found two female-specific alleles, designated W-b (151 bp) and W-d (129 bp). We previously found that 'Dongkui', a female cultivar, could produce viable pollen (we refer to such plants 'Dongkui-male') and serve as the paternal parent in crosses. The genotypes of the MrFT2_BD-SEX marker were W-b/Z in 'Biqi' and W-d/Z in 'Dongkui-male'. The progeny of a cross between these parents produced a 3:1 female (W-) to male (ZZ) ratio and the expected 1:1:1:1 ratio of W-b/W-d: W-b/Z: W-d/Z: Z/Z. In addition, the flowering and fruiting phenotypes of all the F1 progeny fit their genotypes. Our results confirm the existence of ZW sex determination and show that the female phenotype is controlled by a single dominant locus (W) in a small genomic region (59 kb and less than 3.3 cM). Furthermore, we have produced a homozygous "super female" (WW) that should produce all-female offspring in the F2 generation, providing a foundation for commercial use and presenting great potential for use in modern breeding programs.

Genome-wide identification and analysis of the MADS-box gene family and its potential role in fruit development and ripening in red bayberry (Morella rubra).

The MADS-box gene family encodes transcription factors and plays an important role in plant growth and the development of flower and fruit. A perennial dioecious plant, the red bayberry genome has been published recently, providing the opportunity to analyze the MADS-box gene family and its role in fruit development and ripening. Here, we identified 54 MADS-box genes in the red bayberry genome, and classified them into two types based on phylogenetic analysis. Thirteen Type I MADS-box genes were subdivided into three subfamilies and 41 Type II MADS-box genes into 13 subfamilies. A total of 46 MADS-box genes were distributed across eight red bayberry chromosomes, and the other eight genes were located on the unmapped scaffolds. Transcriptome analysis suggested that the expression of most Type II genes was higher than Type I in five female tissues. Moreover, 26 MADS-box genes were expressed during red bayberry fruit development and ten of them showed high expression. qRT-PCR showed that the expression of MrMADS01 (SEP, MIKCC), with differences between the pale pink and red varieties, increased significantly at the final ripening stage, suggesting it may participate in ripening as positive regulator and related to anthocyanin biosynthesis. These results provide some clues for future study of MADS-box genes in red bayberry, especially in ripening process.

Influences of cross pollination on pollen tube growth and fruit set in Zuili plums (Prunus salicina).

Zuili plum (Prunus salicina L.) trees usually set fruit poorly, although they produce high quality fruit. To elucidate the causes of the poor fruit set, pollen tube growth into pistils and fruit set percentage were investigated after cross-, self- and open-pollination. Ovule development in Zuili pistils was also investigated. Pollen tube penetration into the ovules via the obturator and micropyle was best when Zuili pistils were pollinated by cv. Black Amber (P. domestica) pollen grains, although cross-pollinations with Hongxinli and Miili (P. salicina) pollen were more effective than self- and open-pollination. The fruit set percentage was also highest in pistils pollinated with Black Amber pollen grains. Morphological observation of Zuili pistils revealed that the trees produce "double pistils", developing two ovaries from a basal pistil, at a rate as high as 28%. In such abnormal pistils, most ovules were lacking an embryo sac or were entirely degenerated. The percentage of normally developed ovules was 24.3% and 8.9% in normal and double pistils, respectively. From these results, we conclude that the main causes of poor fruit set of Zuili plums are a lack of effective cross-pollination and the production of high percentages of double pistils in which normally developed ovules are scarcely formed.

Effects of post-harvest light conditions on quality and aromatic volatile formation in 'Hakuho' peach (Prunus persica Batsch) fruits.

The effect of light condition during post-harvest storage on fruit quality of 'Hakuho' peach (Prunus persica Batsch) was examined. Fruits were harvested at the immature stage (7 d before the tree-ripening stage) and firm-ripe (3 d before the tree-ripening) stage and stored at 25 degrees C under light (ca 80 micromol m(-2) s(-1) at the fruit top by a fluorescent lamp) and in darkness. The light and dark conditions did not significantly influence the ethylene production rate except for the fully ripened fruits harvested at firm-ripe stage and stored under light. However, no difference in fruit firmness was detected among treatments at full-ripe stage. The skin anthocyanin content increased significantly during storage under light. Total soluble solid (TSS) content of juice at the full ripe stage was not affected significantly by the storage condition, although titratable acidity (TA) in immature harvested fruits decreased more quickly during storage under light compared with those stored in darkness. Dark storage limited the decrease in juice asparagine to some extent. Aromatic lactones, such as gamma-decalactone and gamma-dodecalactone, both in skin and in flesh tissues increased more rapidly when the fruits were stored under a light condition, irrespective of fruit harvest stage. From these results, we conclude that fruit storage under a light condition is better for fruit quality of the 'Hakuho' peaches than storage in darkness.

Effects of fruit pre-harvest bagging on fruit quality of peach (Prunus persica Batsch cv. Hujingmilu).

Experiments were conducted to test the effects of pre-harvest bagging on fruit ripening and quality of peach (Prunus persica Batsch cv. Hujingmilu). Young fruits, at 50 days after full bloom (DAFB), were covered with bags made of single-, double-, and triple-layers of orange paper bag with 27.0%, 13.9% and 8.2% sunlight transmission, respectively. Ethylene production and respiration rate were measured, and fruit quality was analyzed at 111, 114, 117, 120 (firm-ripe stage) and 124 DAFB (full-ripe stage). Single- and triple-layer bagged fruits had higher ethylene production rates than double-layer bagged and un-bagged fruits. The skin of un-bagged fruit had higher brightness (L-value) but smaller hue angle (h degrees) at the full-ripe stage compared with that of bagged fruit. Flesh firmness of un-bagged fruit was higher than that of bagged fruit until the firm-ripe stage, although triple-layer bagged fruit had higher firmness than un-bagged fruit at the full-ripe stage. Total soluble solids in juice of single-layer bagged fruit were a little higher than those of other treated fruits at the full-ripe stage. Single-layer bagged fruit showed the highest level of gamma-decalactone, a main characteristic aroma of peach and total lactones at the firm-ripe and full-ripe stages. It was concluded that 'Hujingmilu' peach had high quality with abundant aromas when the fruits were bagged with single-layer orange paper bags at 50 DAFB. The biosynthesis of gamma-decalactone and other aromas may be affected by light to some extent.

Changes in aroma volatile compounds and ethylene production during "Hujingmilu" peach (Prunus persica L.) fruit development.

Changes in ethylene production rate, respiration rate, and aroma volatile composition was determined from the immature to mature stage in "Hujingmilu" peach (Prunus presica) fruit. Flesh firmness, fruit size, ground color of skin, and total soluble solids (TSS), titratable acidity (TA) in juice were analyzed, and ACC, ACC synthase and ACC oxidase activities were measured. With the increase of fruit maturity, fruit size, L* value, and TSS and major reducing sugar contents increased, whereas fruit firmness, skin hue angle (h degrees ), and juice TA and organic acid contents decreased. In immature fruits, a C(6)-aldehydes (trans-2-hexenal) and C(6)-alcohols (cis-3-hexenol) were the major components, corresponding to the low ethylene production and high respiration rates. With increasing of fruit maturity, the C(6)-C(12) lactonic compounds, particularly gamma- and delta-lactones became the dominant volatile constituents, which increased significantly at the climacteric stage. These facts suggest that ethylene production may be involved in the regulation of lactones production in maturing peach fruit. The activity of ACC synthase in the mesocarp declined during the climacteric rise of ethylene. ACC level and ACC oxidase activity in mesocarp were consistent with the onset of ethylene production in fruit. Major aroma volatiles are thought to form in conjunction with the initiation of climacteric rises in respiration and ethylene production. Detection of changes in aroma volatile compound synthesis rate in peach may be useful as index of maturity reflecting the current physiological stage of peach fruit development.

[Soil respiration and carbon balance in wheat field under conservation tillage].

In order to study the characteristics of carbon sources and sinks in the winter wheat farmland ecosystem in southwest hilly region of China, the LI6400-09 respiratory chamber was adopted in the experiment conducted in the experimental field in Southwest University in Chongqing. The soil respiration and plant growth dynamics were analyzed during the growth period of wheat in the triple intercropping system of wheat-maize-soybean. Four treatments including T (traditional tillage), R (ridge tillage), TS (traditional tillage + straw mulching), and RS (ridge tillage + straw mulching) were designed. Root biomass regression (RR) and root exclusion (RE) were used to compare the contribution of root respiration to total soil respiration. The results showed that the average soil respiration rate was 1.71 micromol x (m2 x s)(-1) with a variation of 0.62-2.91 micromol x (m2 x s)(-1). Significant differences in soil respiration rate were detected among different treatments. The average soil respiration rate of T, R, TS and RS were 1.29, 1.59, 1.99 and 1.96 micromol x (m2 x s)(-1), respectively. R treatment did not increase the soil respiration rate significantly until the jointing stage. Straw mulching treatment significantly increased soil respiration, with a steadily high rate during the whole growth period. During the 169 days of growth, the total soil respiration was 2 266.82, 2799.52, 3 483.73 and 3 443.89 kg x hm(-2) while the cumulative aboveground biomasses were 51 800.84, 59 563.20, 66 015.37 and 7 1331.63 kg x hm(-2). Compared with the control, the yield of R, TS and RS increased by 14.99%, 27.44% and 37.70%, respectively. The contribution of root respiration to total soil respiration was 47.05% by RBR, while it was 53.97% by RE. In the early growth period, the carbon source was weak. The capacity of carbon sink started to increase at the jointing stage and reached the maximum during the filling stage. The carbon budget of wheat field was 5 924.512, 6743.807, 8350.741, 8 876.115 kg x hm(-2), respectively. The results indicated that ridge tillage and straw mulching conservation tillage significantly improved the carbon sink in the wheat farmland ecosystem.

Development of 107 SSR markers from whole genome shotgun sequences of Chinese bayberry (Myrica rubra) and their application in seedling identification.

Chinese bayberry (Myrica rubra Sieb. et Zucc.) is one of the important subtropical fruit crops native to the South of China and Asian countries. In this study, 107 novel simple sequence repeat (SSR) molecular markers, a powerful tool for genetic diversity studies, cultivar identification, and linkage map construction, were developed and characterized from whole genome shotgun sequences. M13 tailing for forward primers was applied as a simple method in different situations. In total, 828 alleles across 45 accessions were detected, with an average of 8 alleles per locus. The number of effective alleles ranged from 1.22 to 10.41 with an average of 4.08. The polymorphic information content (PIC) varied from 0.13 to 0.89, with an average of 0.63. Moreover, these markers could also be amplified in their related species Myrica cerifera (syn. Morella cerifera) and Myrica adenophora. Seventy-eight SSR markers can be used to produce a genetic map of a cross between 'Biqi' and 'Dongkui'. A neighbor-joining (NJ) tree was constructed to assess the genetic relationships among accessions, and the elite accessions 'Y2010-70', 'Y2012-140', and 'Y2012-145', were characterized as potential new genotypes for cultivation.

Anatomical studies of ovule development in the post-bloom pistils of the 'Zuili' plum (Prunus salicina Lindl.).

In order to investigate the cause of poor fruit set in 'Zuili' plums, anatomical examinations of post-bloom pistils were conducted and the dates of young fruit drop were recorded during the growing seasons of 2008 and 2009. Pistils of cv. 'Black Amber' were also examined as an abundant setting control. Two major dropping periods were detected in 'Zuili': one during the first 5 d after full bloom (DAF) and another between 10 and 17 DAF. Anatomical analyses of the pistils at the full bloom stage revealed that half of the ovules had not developed embryos, which may have caused their early drop. In most dropped pistils collected at 17 DAF, the micropyle had not been penetrated by a pollen tube, indicating that they were not fertilized. 'Zuili' ovules initiated embryo division at 10-12 DAF, although thereafter embryo development was retarded when compared to the rates observed in 'Black Amber'. Ovule fertilization failure and inactive embryo development after ovule fertilization may be the major causes of the later fruit drop observed in 'Zuili' plum trees.