Cryopreservation of Schisandra chinensis (Turcz.) Baill callus and subsequent plant regeneration.

Cryopreservation has been proven significance as a technique for promising the long-term conservation of plant germplasms. This study aimed to establish a cryopreservation protocol for calli of Schisandra chinensis (Turcz.) Baill, and to explore the effects of different process parameters on callus viability. Effects of desiccation duration, cryoprotectants and cryopreservation methods, thawing temperature, and post-culture conditions on the viability of cryopreserved calli were assessed. Among different cryoprotectants and freezing procedures, the highest survival was recorded when the water content of callus after 30 min desiccation was 57.3%, were loaded into a cryoprotectant containing 10% ethylene glycol, 8% glucose, and 10% DMSO, and frozen slowly (-1°C/min). Rapid thawing at 40°C for 2 min demonstrated the best recovery of cryopreserved S. chinensis calli. Post-culturing in darkness for one week before transfer to light conditions (under 16 h photoperiod at 36 µmol·m-2·s-1) was beneficial to callus regeneration. Plants regenerated through somatic embryogenesis from cryopreserved calli remained ploidy stable after cryopreservation. The callus cryopreservation procedure established in this study is a promising tool for the conservation of S. chinensis resources.

Karyotype studies on Lycoris radiata populations from China.

Lycoris radiata is an important medicinal and ornamental plant of China. In the present study, somatic chromosome counts and karyotype analyses, which are important aspects of plant phylogeny and evolution, were performed in 466 individuals from 25 L. radiata populations by root tip squash method. Chromosome counts revealed that 10 populations were diploid (2n = 2x = 22) and 15 were triploid (2n = 3x = 33). Except for one diploid population containing some triploid plants, the remaining 24 populations showed a single cytotype. Karyotype analysis showed that the karyotypes of L. radiata varied in different populations and even within the same population. However, based on the Stebbins' system, the karyotype of all the populations could be classified in 4A classes. The cluster analysis and ordination methods demonstrated that the L. radiata populations grouped in two major clusters. Previous research has shown that the triploid strain of L. radiata is a genetically identical species. However, the cluster analysis revealed that the triploid strains clustered in two groups instead of one, which indicates that these strains may not be identical species, genetically. This study is expected to improve the understanding of the genetic diversity in L. radiata and provide a basis for future studies on species differentiation, speciation, and taxonomy.

Discovery and characterization of a first set of polymorphic microsatellite markers in Siganus oramin.

Nine microsatellite DNA markers were developed and characterized for Siganus oramin by the 5'-anchored polymerase chain reaction technique. A total of 42 alleles were identified in 30 individuals, and the number of alleles per locus ranged from 3 to 7, with an average of 4.7. The observed and expected heterozygosity per locus ranged from 0.5333 to 1.0000 and from 0.5254 to 0.8474, respectively, with an average of 0.7422 and 0.6906, respectively. A significant deviation from the Hardy-Weinberg equilibrium was detected at one microsatellite locus after a Bonferroni's correction (P < 0.0056). No significant linkage disequilibrium was found between any of the pairs of the nine loci. The microsatellite loci developed in this study will improve our understanding of the genetic background of S. oramin.

Effect of wilfortrine on human hepatic cancer HepG2 cell proliferation potential in vitro.

Liver cancers are characterized by high morbidity and mortality owing to few effective drugs for its treatment. Wilfortrine has several pharmacological effects, including an inhibitory effect on liver cancer cell proliferation. However, whether wilfortrine can induce liver cancer cell apoptosis has not been elucidated. We investigated the role of wilfortrine on liver cancer cell HepG2 apoptosis and analyzed its possible mechanisms to provide a theoretical basis for clinical analysis of liver cancer pathogenesis. The liver cancer cell line HepG2 was treated with 40 mM wilfortrine for 48 h. Flow cytometry was applied to detect HepG2 cell apoptosis and cell cycle changes. Western blot was used to analyze Bcl-2 and Bax expression. The HepG2 cell apoptosis rate increased significantly after treatment with wilfortrine, especially the early apoptosis rate (P < 0.05). However, wilfortrine did not change the cell cycle of HepG2 cells. After wilfortrine treatment, Bcl- 2 expression decreased significantly (P < 0.05); on the contrary, Bax expression increased noticeably compared with the control group (P < 0.05). Wilfortrine can induce liver cancer cell HepG2 apoptosis, but with no effect on the cell cycle, mainly by promoting Bax expression and inhibiting anti-apoptotic protein Bcl-2 expression.

Prediction of core cancer genes using a hybrid of feature selection and machine learning methods.

Machine learning techniques are of great importance in the analysis of microarray expression data, and provide a systematic and promising way to predict core cancer genes. In this study, a hybrid strategy was introduced based on machine learning techniques to select a small set of informative genes, which will lead to improving classification accuracy. First feature filtering algorithms were applied to select a set of top-ranked genes, and then hierarchical clustering and collapsing dense clusters were used to select core cancer genes. Through empirical study, our approach is capable of selecting relatively few core cancer genes while making high-accuracy predictions. The biological significance of these genes was evaluated using systems biology analysis. Extensive functional pathway and network analyses have confirmed findings in previous studies and can bring new insights into common cancer mechanisms.

Characteristics of growth traits and their effects on body weight of G1 individuals in the mud crab (Scylla paramamosain).

The mud crab (Scylla paramamosain) is considered a potentially important marine crab species for selective breeding. Here, we first examined sex ratio and differences in 16 growth traits between females and males in a G1 population of S. paramamosain, and we then analyzed the correlation between these growth traits and their effects on body weight (BW). Of these growth traits, nine were significantly different between sexes. In females, the correlation coefficients in all trait pairs ranged from 0.524 to 0.997. The traits carapace length (CL) and distance between lateral spine 2 (DLS2) significantly affected BW directly, with the path coefficients being 1.124 and -0.186, respectively. The determination coefficients of traits CL and DLS2 to BW were 1.263 and 0.035 with the total value being 0.951, indicating that the two traits were the key factors affecting BW. In males, the correlation coefficients in all trait pairs ranged from 0.881 to 0.999. The three traits body height (BH), fixed finger height of the claw (FFHC), and meropodite length of pereopod 2 (MLP2) significantly affected BW directly, with the path coefficients being 0.484, 0.300, and 0.225, respectively. The determination coefficients of traits BH, FFHC and MLP2 to BW were 0.234, 0.090 and 0.051, with the total value being 0.967, indicating that these three traits played a key role in affecting BW. Moreover, we constructed two best-fit linear regression equations, which were Y (BW) = 4.969 X1 (CL) - 0.758 X2 (DLS2) - 140.177 and Y (BW) = 3.806 X1 (BH) + 2.371 X2 (FFHC) + 1.725 X3 (MLP2) - 123.559 in females and males, respectively.

Polymorphisms of KAP6, KAP7, and KAP8 genes in four Chinese sheep breeds.

High glycine-tyrosine proteins (HGTPs), also known as keratin-associated proteins (KAPs), play a key role in the major structures and mechanical properties of wool fiber. Sheep HGTPs consist of three multigene families: KAP6, KAP7, and KAP8 genes. Polymorphisms of these three genes have been proposed to have important effects on wool fiber traits. The aim of the present study was to identify polymorphisms of the KAP6, KAP7, and KAP8 genes in four sheep breeds, including Chinese Merino superfine wool sheep, Hu sheep, a Merino x Hu crossed breed, and Romney sheep. Polymerase chain reaction (PCR) product direct sequencing, PCR-single-strand conformation polymorphism, and cloned sequencing methods were used to find genetic variation and identify polymorphisms in these genes. The Mutation Surveyor v3.97 software was used to analyze the sequences. These methods revealed six different sequences of the KAP6 gene, two different sequences of the KAP7 gene, and five different sequences of the KAP8 gene. Accordingly, three (with frequencies>1%) single nucleotide polymorphisms (SNPs) of the KAP6 gene, one SNP of the KAP7 gene, and five SNPs of the KAP8 gene were detected. Interestingly, some of these sequences were present in only certain sheep breeds, thereby suggesting that these special allele sequences could be used as candidate genes of wool characteristics in further studies.

Polymorphisms of the ATP1A1 gene associated with mastitis in dairy cattle.

Mastitis affects the concentrations of potassium and sodium in milk. Since sodium-potassium adenosine triphosphatase (Na(+), K(+)-ATPase) is critical for maintaining the homeostasis of these two ions, and is involved in cell apoptosis and pathogenesis, we presumed that polymorphism of the ATP1A1 gene, which encodes the bovine Na(+), K(+)-ATPase α1 subunit could be associated with mastitis. The ATP1A1 gene was analyzed in 320 Holstein cows using PCR low ionic strength single-strand conformation polymorphism (PCR-LIS-SSCP) and DNA sequencing methods. A C/A SNP was identified at nucleotide position -15,739 in exon 17 of the ATP1A1 gene, but it did not induce any change in amino acids. We examined a possible association of polymorphism of the ATP1A1 gene with somatic cell score and 305-day milk yields. Individuals with genotype CC in ATP1A1 had significantly lower somatic cell scores and 305-day milk yields than those with genotype CA. We also examined changes in Na(+), K(+)-ATPase activity of red cell membranes. The Na(+), K(+)-ATPase activity was significantly higher in dairy cows with genotype CC compared to the other two genotypes, and the Na(+), K(+)-ATPase activity of the resistant group was significantly higher than that of the susceptible group in dairy cows. We conclude that this polymorphism has potential as a marker for mastitis resistance in dairy cattle.

Regulation of cotton fiber elongation by xyloglucan endotransglycosylase/hydrolase genes.

Ligon lintless mutant (li1li1) with super-short fibers (5-8 mm in length) and its wild type (Li1Li1) with normal fibers (30 mm in length) were used to study the function of xyloglucan endotransglycosylase/hydrolase (XTH) genes during fiber elongation in cotton. Wild-type cotton attained the fiber elongation stage earlier (5 days post-anthesis, DPA), than the Ligon lintless mutant (12 DPA) with a higher fiber elongation velocity of about 1.76 mm/day. Xyloglucan contents in Ligon lintless mutant fibers were 5-fold higher than the wild type during 9-15 DPA. It was also observed that the activity of XTH in wild-type cotton fibers was about 2-fold higher than that of the Ligon lintless mutant with a peak at 12 DPA. DNA blot analysis indicated that the XTH gene in the Ligon lintless mutant and its wild type belonged to a multiple allelic series. However, RNA blot analysis and quantitative real-time PCR exhibited an earlier expression (10 DPA) of XTH in wild type as compared to delayed (15 DPA) expression in the Ligon lintless mutant. The study also revealed that 9-15 DPA might be a key phase for upregulation of fiber elongation via increasing XTH activity. Higher XTH activity can cleave down the xyloglucan-cellulose chains thus loosening fiber cell wall and promoting fiber cell elongation in wild type as compared to its mutant.

Association of ATP1A1 gene polymorphism with heat tolerance traits in dairy cattle.

Heat stress produces oxidative stress and affects the alternation of plasma K(+) and Na(+). Since Na(+),K(+)-ATPase is sensitive to oxidative stress and critical for maintaining the homeostasis of these two ions, we examined the genetic polymorphism of the ATP1A1 gene in 160 Holstein cows using polymerase chain reaction low ionic strength single-strand conformation polymorphism and DNA sequencing methods. G to A at position -14103 in exon 14 and C to T at position -14242 in intron 14 of the bovine ATP1A1 gene were identified, but the former single nucleotide polymorphism was silent with respect to the amino acid sequence of the protein. However, we found significant correlations between ATP1A1 gene polymorphism and the coefficient of heat tolerance (P<0.01) and with respiratory rate (P<0.01). Genotype AC was the most favorable genotype for heat tolerance. This polymorphism site has potential as a genetic marker for heat tolerance traits in dairy cattle breeding.