A high-density genetic map developed by specific-locus amplified fragment (SLAF) sequencing and identification of a locus controlling anthocyanin pigmentation in stalk of Zicaitai (Brassica rapa L. ssp. chinensis var. purpurea).

BACKGROUND: Caixin and Zicaitai (Brassica rapa) belong to Southern and Central China respectively. Zicaitai contains high amount of anthocyanin in leaf and stalk resulting to the purple color. Stalk is the major edible part and stalk color is an economically important trait for the two vegetables. The aim of this study is to construct a high density genetic map using the specific length amplified fragment sequencing (SLAF-seq) technique to explore genetic basis for anthocyanin pigmentation traits via quantitative trait loci (QTL) mapping. RESULTS: We constructed a high generation linkage map with a mapping panel of F2 populations derived from 150 individuals of parental lines "Xianghongtai 01" and "Yinong 50D" with purple and green stalk respectively. The map was constructed containing 4253 loci, representing 10,940 single nucleotide polymorphism (SNP) markers spanning 1030.04 centiMorgans (cM) over 10 linkage groups (LGs), with an average distance between markers of 0.27 cM. Quantitative trait loci (QTL) analysis revealed that a major locus on chromosome 7 and 4 minor QTLs explaining 2.69-61.21% of phenotypic variation (PVE) were strongly responsible for variation in stalk color trait. Bioinformatics analysis of the major locus identified 62 protein-coding genes. Among the major locus, there were no biosynthetic genes related to anthocyanin. However, there were several transcription factors like helix-loop-helix (bHLH) bHLH, MYB in the locus. Seven predicted candidate genes were selected for the transcription level analysis. Only bHLH49 transcription factor, was significantly higher expressed in both stalks and young leaves of Xianghongtai01 than Yinong50D. An insertion and deletion (InDel) marker developed from deletion/insertion in the promoter region of bHLH49 showed significant correlation with the stalk color trait in the F2 population. CONCLUSION: Using the constructed high-qualified linkage map, this study successfully identified QTLs for stalk color trait. The identified valuable markers and candidate genes for anthocyanin accumulation in stalk will provide useful information for molecular regulation of anthocyanin biosynthesis. Overall our findings will lay a foundation for functional gene cloning, marker-assisted selection (MAS) and molecular breeding of important economic traits in B. rapa.

Effect of Modified Wuzi Yanzong Pill () on Tip60-Mediated Apoptosis in Testis of Male Rats after Microwave Radiation.

OBJECTIVE: To investigate the effect of a modified Wuzi Yanzong Pill (, WZYZP) on the male rats' testis after microwave radiation, as well as its potential mechanism. METHODS: Forty-five male rats were randomly assigned to three groups: the control group, the radiation group, and the WZYZP group. The rats in the radiation group and WZYZP group were exposed to microwave radiation for 15 min once, while the rats in the control group were not exposed to any radiation. The rats in the WZYZP group were given a modified of WZYZP by gavage daily for 7 days. Apoptosis in the testis was evaluated using terminal-deoxynucleoitidyl transferase mediated nick end labeling (TUNEL) assay. Histopathological alterations of the testis were observed by haematoxylin-eosin (HE) staining. Tat-interactive protein, 60kD (Tip60) and p53 expressions were determined by Western blotting. RESULTS: The apoptosis index (AI) in the radiation group was higher than that of the WZYZP group and control group on day 1 (D1), day 7 (D7) day 14 (D14) after radiation (P<0.05). The seminiferous tubules were of normal morphology in the control group. In the radiation group, the partial seminiferous tubules were collapsed, basement membranes of the seminiferous epithelia became detached. WZYZP could restore the morphological changes. There was no expression of Tip60 among the three groups on D7 and D14. The expression of p53 was higher in the radiation group than in the control group (P<0.05). WZYZP could down-regulate the rising p53 induced by radiation on D7 and D14 (P<0.05). CONCLUSION: A modified WZYZP may affect germ cells, and its protective effects may partly result from its ability to intervene in Tip60 mediated apoptosis.

A high-density linkage map and QTL mapping of fruit-related traits in pumpkin (Cucurbita moschata Duch.).

Pumpkin (Cucurbita moschata) is an economically worldwide crop. Few quantitative trait loci (QTLs) were reported previously due to the lack of genomic and genetic resources. In this study, a high-density linkage map of C. moschata was structured by double-digest restriction site-associated DNA sequencing, using 200 F2 individuals of CMO-1 × CMO-97. By filtering 74,899 SNPs, a total of 3,470 high quality SNP markers were assigned to the map spanning a total genetic distance of 3087.03 cM on 20 linkage groups (LGs) with an average genetic distance of 0.89 cM. Based on this map, both pericarp color and strip were fined mapped to a novel single locus on LG8 in the same region of 0.31 cM with phenotypic variance explained (PVE) of 93.6% and 90.2%, respectively. QTL analysis was also performed on carotenoids, sugars, tuberculate fruit, fruit diameter, thickness and chamber width with a total of 12 traits. 29 QTLs distributed in 9 LGs were detected with PVE from 9.6% to 28.6%. It was the first high-density linkage SNP map for C. moschata which was proved to be a valuable tool for gene or QTL mapping. This information will serve as significant basis for map-based gene cloning, draft genome assembling and molecular breeding.

[Yijingfang for the treatment of asthenospermia: A randomized controlled clinical trial].

OBJECTIVE: To investigate the clinical therapeutic effects of Yijingfang, a Chinese medicinal liquid, on asthenospermia. METHODS: We randomly divided 450 asthenospermia patients into a treatment group (n = 300) and a control group (n = 150), the former treated with Yijingfang once half a dose, bid, and the latter with Wuziyanzong Pills (9 g, bid) + L-carnitine oral liquid (10 ml, bid), both for 3 months. Before and at 1, 2, and 3 months after medication, we compared the semen volume, sperm concentration, percentages of progressively motile sperm (PMS) and total motile sperm (TMS), and semen liquefaction time between the two groups of patients. RESULTS: No statistically significant difference was observed in the semen parameters between the treatment and control groups before medication (P >0.05). In comparison with the baseline, the treatment group showed significant differences at 1, 2, and 3 months after medication in sperm concentration (ï¼»35.96 ± 8.50ï¼½ vs ï¼»49.66 ± 10.91ï¼½, ï¼»55.21 ± 11.46ï¼½, ï¼»74.90 ± 13.07ï¼½ ×106/ml, P <0.01), PMS (ï¼»19.72 ± 2.06ï¼½ vs ï¼»23.81 ± 2.56ï¼½, ï¼»26.12 ± 2.34ï¼½, and ï¼»32.17 ± 1.62ï¼½ %, P <0.01) and TMS (ï¼»28.86 ± 2.70ï¼½ vs ï¼»34.17 ± 3.43ï¼½, ï¼»36.59 ± 3.36ï¼½, and ï¼»47.08 ± 2.97ï¼½ %, P <0.01), but not in the semen volume (ï¼»3.35 ± 0.99ï¼½ vs ï¼»3.15 ± 1.06ï¼½, ï¼»3.12 ± 0.90ï¼½, and ï¼»3.27 ± 0.78ï¼½ ml, P >0.05) or semen liquefaction time (ï¼»32.31 ± 8.15ï¼½ vs ï¼»31.68 ± 3.14ï¼½, ï¼»30.38 ± 3.44ï¼½, and ï¼»30.86 ± 2.42ï¼½ min, P >0.05); the control group exhibited similar results at the three time points in sperm concentration (ï¼»36.85 ± 6.88ï¼½ vs ï¼»40.53 ± 8.32ï¼½, ï¼»47.51 ± 12.73ï¼½, and ï¼»56.14 ± 11.98ï¼½ ×106/ml, P <0.01), PMS (ï¼»20.26 ± 2.73ï¼½ vs ï¼»25.17 ± 2.64ï¼½, ï¼»27.23 ± 2.25ï¼½, and ï¼»31.89±2.27ï¼½ %, P <0.01), and TMS (ï¼»30.03 ± 2.67ï¼½ vs ï¼»33.89±2.26ï¼½, ï¼»37.38±4.79ï¼½, and ï¼»40.35±3.06ï¼½ %, P <0.01), but not in the semen volume (ï¼»3.03 ± 1.09ï¼½ vs ï¼»3.16±1.78ï¼½, ï¼»3.15±0.96ï¼½, and ï¼»3.12±0.65ï¼½ ml, P >0.05) or semen liquefaction time (ï¼»30.25 ± 5.20ï¼½ vs ï¼»29.36±4.25ï¼½, ï¼»28.21±3.26ï¼½, and ï¼»28.33±3.59ï¼½ min, P >0.05). There were statistically significant differences between the treatment and control groups in the increase rates of sperm concentration and TMS after medication (P <0.01) but not in that of PMS (P >0.05). CONCLUSIONS: Yijingfang is an effective drug for the treatment of asthenospermia, which can regulate the spermatogenesis, increase the percentage of PMS, and improve the total sperm motility of the patients.

[Triple acupuncture at the Qugu acupoint as an adjunctive therapy for type-â ¢ chronic prostatitis: Analysis of short- and long-term clinical effects].

OBJECTIVE: To investigate the short- and long-term effects of triple acupuncture at the Qugu acupoint as an adjunctive therapy on type-Ⅲ chronic prostatitis / chronic pelvic pain syndrome (CP/CPPS). METHODS: We equally randomized 90 CP/CPPS patients into a control and a treatment group, both treated with Levofloxacin Mesylate Tablets (0.5 g, tid) + Terazosin Hydrochloride Capsules (2 mg qd) for 4 weeks, while the latter group by triple acupuncture at the Qugu acupoint as an adjunctive therapy twice a week at the same time. Then, we followed up all the patients for 4 weeks, recorded the cases, time and rate of recurrence, obtained the scores in National Institutes of Health Chronic Prostatitis Symptom Index (NIH-CPSI), quality of life (QoL) and Zung Depression Scale (ZDS), and compared them between the two groups of patients. RESULTS: Compared with the controls, the patients of the treatment group showed significantly decreased NIH-CPSI scores in pain (8.6 ± 2.12 vs 6.2 ± 2.25, P <0.05), micturition (5.8 ± 1.22 vs 3.1 ± 1.10, P <0.05), and QoL (6.0 ± 1.33 vs 3.4 ± 1.71, P <0.05) and ZDS score as well (43.9 ± 4.53 vs 33.6 ± 3.20, P <0.01). The recurrence rate was markedly lower while the recurrence time remarkably longer in the treatment than in the control group (15.56 vs 35.56% and ï¼»20.0 ± 2.72ï¼½ vs ï¼»12.5 ± 3.47ï¼½ d, P <0.05). CONCLUSIONS: As an adjunctive therapy, triple acupuncture at the Qugu acupoint can evidently ameliorate the clinical symptoms, enhance the curative effect of antibacterials, reduce the recurrence rate, and prolong the recurrence time in the treatment of CP/CPPS.

[Regulatory effect of yijing fang on adenine-induced infertility in rat models: a pharmacodynamic study].

OBJECTIVE: To investigate the regulatory effect of Yijing Fang (YJF) on adenine-induced infertility in rats with kidney deficiency. METHODS: Sixty healthy Wistar male rats, aged 1.5 mo and weighing (180 +/- 10) g, were normally fed for a week, and then divided into five groups of equal number (blank control, infertile model, high-dose YJF, mid-dose YJF, and low-dose YJF) according to the body weight of the rats. The models were made by intragastric administration of 500 mg/ml adenine in gum arabic solution in the ratio of 1:10 at the dose of 1 ml per 100 g body weight per day for 10 days. YJF was given at 3.38 g, 1.69 g and 0.85 g per 100 g body weight per day to the rats in the high-, mid- and low-dose groups, respectively. After 48 days of treatment, we observed kidney deficiency-related changes in sperm concentration and motility, the levels of testosterone (T) and other hormones and the volumes of the testis, epididymis, seminal vesicle and prostate, and compared the indexes among different groups. RESULTS: YJF exhibited a significant regulatory effect on sperm concentration and motility, the T level and the indexes of the gonad and other accessory glands in the model rats (P < 0.05). After 48 days of treatment, sperm concentrations were (87.85 +/- 28.44), (7.11 +/- 2.15), (35.98 +/- 14.04), (32.65 +/- 11.80) and (33.51 +/- 13.26) x 10(6)/ml in the blank control, infertile model, high-dose YJF, mid-dose YJF, and low-dose YJF groups, respectively; sperm motilities were (52.79 +/- 16.43), (31.14 +/- 3.07), (45.88 +/- 16.97), (51.56 +/- 13.35) and (49.53 +/- 10.16)%; the T levels were (194.07 +/- 40.29), (61.27 +/- 13.70), (121.87 +/- 24.35), (127.44 +/- 19.38) and (127.81 +/- 20.28) nmol/L; the luteinizing hormone (LH) levels were (7.017 +/- 0.269), (6.117 +/- 0.894), (7.060 +/- 0.871), (7.156 +/- 0.937) and (6.967 +/- 0.778) IU/L; the testis volumes were (3.775 +/- 0.183), (2.865 +/- 0.258), (3.236 +/- 0.058), (3.457 +/- 0.066) and (3.398 +/- 0.091) g; the epididymis volumes were (1.119 +/- 0.116), (0.833 +/- 0.226), (1.124 +/- 0.104), (1.132 +/- 0.107) and (1.114 +/- 0.106) g; the prostate volumes were (176.75 +/- 427.09), (131.67 +/- 39.45), (178.70 +/- 37.97), (180.11 +/- 37.39) and (179.00 +/- 35.42) mg; and the body weights were (188.50 +/- 7.12), (189.92 +/- 6.67), (187.42 +/- 5.47), (189.17 +/- 6.19) and (188.75 +/- 6.12) g. Testis histopathology showed obvious injuries in the infertile models and different degrees of improvement in the three YJF groups, most evidently in the mid-dose group. CONCLUSION: Yifing Fang had an evident therapeutic effect on kidney deficiency-related infertility in adenine-induced rat models.

[Effect of PP333 treatment on seed yield and root of Flemingia philippinensis].

OBJECTIVE: To study the effect of PP333 treatment on seed yield,medicinal materials yield and quality of Flemingia Philippinensis. METHODS: The experiment was in split plot design for PP333 in main plot and in planting density subplot. RESULTS: The seed yield of 600 mg/L (PP333) + 60 plants/m2 (planting density) was the highest. The medicinal materials yield of 600 mg/L (PP333) + 90 plants/m2 (planting density) was the highest. The quality of 900 mg/L( PP333) + 60 plants/m2 (planting density) was the best. CONCLUSION: The appropriate density of PP333 and planting can improve seed yield,medicinal materials yield and quality of Flemingia Philippinensis.

[Yijingfang increases the expression of CatSper1 in mice].

OBJECTIVE: To observe the effects of Yijingfang on CatSper1 in the mouse model of cyclophosphamide-induced oligoasthenospermia. METHODS: Forty Kunming male mice were randomly divided into a control group (CG), a model group (MG), a small-dose Yijingfang group (SG), and a large-dose Yijingfang group (LG). The mice of CG were intraperitoneally injected with normal saline at 60 mg/kg once a day, while those of MG, SG and LG with cyclophosphamide, all for 5 days. During the next 34 days, the mice of SG and LG received intragastric administration of Yijingfang once a day, the former at a dose 2 times and the latter 5 times that of human routine usage, those of MG given the same volume of normal saline, and CG normally fed. At 35 days, we measured the sperm count, percentages of grades a + b and a + b + c sperm, and the expression of CatSper1 in the epididymal sperm of the mice. RESULTS: The sperm counts of CG, MG, SG and LG were (5.20 +/- 1.34), (1.73 +/- 0.03), (2.08 +/- 0.01) and (3.31 +/- 0.56) x 10(6)/ml, respectively, significantly lower in MG than in CG (P < 0.05), but higher in LG than in MG (P < 0.05). The grade a + b sperm constituted (14.49 +/- 0.30), (6.64 +/- 1.88), (11.99 +/- 1.01) and (19.40 +/- 3.13)% in CG, MG, SG and LG, respectively, remarkably lower in MG than in CG (P < 0.05) but higher in LG than in MG (P < 0.05); the grade a + b + c sperm accounted for (68.39 +/- 15.13), (39.96 +/- 4.89), (62.28 +/- 4.43) and (73.61 +/- 5.05)%, respectively, obviously lower in MG than in CG (P < 0.05) but higher in LG than in MG (P < 0.05); the CatSper1 expressions were 0.76 +/- 0.05, 0.73 +/- 0.03, 0.75 +/- 0.12 and 0.85 +/- 0.04, respectively, markedly higher in LG than in MG (P < 0.05). CONCLUSION: Intraperitoneal injection of cyclophosphamide decreases the sperm count, percentages of grades a + b and a + b + c sperm, and the expression of CatSper1 in mice, while large-dose Yijingfang can increase the above parameters, and hence contributes to the treatment of oligoasthenospermia.

[Longbixiao capsule reduces expressions of TGF-beta1 and Smoothelin in human prostatic stromal cells cultured in vitro].

OBJECTIVE: To investigate the effects of the Chinese herbal medicine of Longbixiao (LBX) Capsule on the expressions of TGF-beta1 and Smoothelin in human prostatic stromal cells cultured in vitro. METHODS: Blood serum medicated with LBX was incubated with the stromal cells isolated from men with benign prostatic hyperplasia (BPH) and cultured in vitro. The mRNA expression levels of TGF-beta1 and Smoothelin were detected by real-time RT-PCR and other relevant techniques. RESULTS: In the high and low concentration groups, the gene relative expressions of TGF-beta1 were (0.158 +/- 0.020) and (0.169 +/- 0.020) , while those of Smoothelin were (0.035 +/- 0.007) and (0.036 +/- 0.007) respectively, both significantly decreased in comparison with the control group(P < 0.01). CONCLUSION: LBX reduces the mRNA expressions of TGF-beta1 and Smoothelin in human prostatic stromal cells and can be used in the treatment of BPH.