Long-term in-vitro culture and subculture of the hemocytes of swimming crab Portunus trituberculatus

Crab cell line,especially continuous crab cell line,can provide us a useful tool for studies on the virology,immunology,and molecular biology of crabs.However,no continuous crab cell line has been available due to the lacking of suitable medium and the occurrence of mitosis-arrest.In this study,long-term in vitro culture conditions for both two-(2D)and three-dimensions(3D)were successfully developed for the circulating hemocytes of swimming crab Portunus trituberculatus,designated as PTH cells.In 2D culture,a novel crab basic medium in osmolarity of 990–1100 mOsm/kg was optimized for the first time,which is different from Leibovitz's L-15 medium in mainly the components of amino acids,containing double strengths of the contents of free amino acid mixture in the crab serum.Then an optimal crab growth medium was developed by supplementing 5%fetal bovine serum,50-g/L yeast extract powder,20-μg/L basic fibroblast growth factor and epidermal growth factor into the optimal crab basic medium,and found that it could support a long-term survival of PTH cells in a healthy monolayer up to 347 days and partially break through the mitosis-arrest of crab cells evidenced by the obvious increase of proliferating potential detected in the 10-d primarily cultured PTH cells.These 2D cultured PTH cells could be successfully sub-cultured for 11 times by physical flushing method and well cryopreserved in liquid nitrogen.In 3D culture,using the same crab growth medium,the PTH cell aggregates could be easily formed and healthily maintained on the surface of solidified Matrigel or in the ultra-low-attachment plate with a survival rate of 50%–60%on Day 103.This work largely improved the primary culture and subculture of crab cells and will facilitate the establishment of continuous crab cell line.

6 Kinds of Isozymes after Long-term Subculture of Emmenopterys henryi Oliv.

[Objective] The aim was to study the changes of zymography in 6 kinds of isozymes after long-term subculture of Emmenopterys henryi Oliv.. [Method] Non-denaturing polyacrylamide gel electrophoresis was used to analyze isozyme patterns such as esterase (EST),acid phosphatase (ACP),ATP enzyme (ATPase),amylase (AMY),superoxide dismutase (SOD) and peroxidase (POD) in long-term subculture callus of Emmenopterys henryi Oliv. [Result] The research showed that there were differences among the 6 kinds of isozymes in embryogenic callus and non-embryogenic callus of Emmenopterys henryi Oliv.,and both levels could be taken as the basis for identification,the EST,ACP,and POD of non-embryogenic callus were significantly higher than embryogenic callus. The browning of non-embryogenic callus was non-level in the AMY,SOD and POD isozymes when was compared with normal non-embryogenic callus,while the EST,ACP and ATPase isozymes decreased; When the browning of embryogenic callus was contrasted with normal embryogenic callus,EST isozyme increased and the other 5 kinds of enzymes decreased. [Conclusion] The study provided theoretical basis for research morphological difference and browning of long-term tissue culture of Emmenopterys henryi Oliv..

Effects of in vitro subculture on the physiological characteristics of adventitious root formation in microshoots of Castanea mollissima cv.'yanshanhong'

Microshoots of Castanea mollissima cv.＇yanshanhong＇ in vitro acquired an enhanced rooting capability with increasing numbers of subculture.In this study,we investigated the effect of successive subculture on adventitious root formation in vitro by the determination of the endogenous hormone level and the enzyme activity.The levels of indoleacetic acid（IAA）,abscisic acid（ABA）,cytokinins（CTK） and gibberellic acid（GA3） were determined by high performance liquid chromatography（HPLC）,and the activities of indoleacetic acid oxidase（IAAO）,peroxidase oxidase（POD）,and polyphenol oxidase（PPO） were measured by ultraviolet-spectrophotometer assay after the induction of rooting at 2nd,4th,6th and 8th subculture.The relationships between physiological characteristics and subculture numbers or rooting rate were as follows：The levels of endogenous IAA in microshoots gradually increased,and endogenous levels of ABA,CTK and GA3 in microshoots decreased slightly after serial subcultures.The level of IAA was highly correlated with subculture numbers and rooting rates.The ratios of IAA/ABA and IAA/CTK both acutely raised with increasing rooting rate during successive subcultures and had high correlations with rooting rate.The activity of IAAO and POD are significantly negatively related with subculture numbers,and the activity of PPO increased after subcultures.

Cultures and Subcultures

A cross-cultural analysis of an extract of the book Lost in Translation is provided in the essay. Definitions of culture and subculture are discussed and different theories from Hofstede and Schwartz are referred to in order to validate the analysis.

The Analysis on the Influence of Amarican Subculture upon American College Slang

The thesis is just trying to analyze American college slang from sub-cultural perspective on the basis of combining linguistic knowledge,the knowledge of American slang and American college slang with the knowledge of subculture,which is helpful for English learners to learn American English better.

A Study of the Subculture of Human Endothelial Cell From Umbilical Vein

Human endothelial cells derived from umbilical vein could besubcultured and survive 25 passage (double time 20～25 hours, 60～75 cumula-tive population doublings). An observation was performed with light mi-croscopy, electron microscopy and immunofluorescence microscopy using specificantiserum against factor Ⅷrelated antigen. It identified that the cultured cellswere endothelial cells. Medium RPMI-1640 supplemented with 20% humanserum, endothelial cell growth factor 200μg/ml, heparin 100μg/ml and gelatincoated flasks were very important conditions for long-term culture of humanendothelial cell.

The Potential of Subculture Helicoverpa armigera Nuclear Polyhedrosis Virus as an Alternate Synthetic Insecticides to Control Insect Pests in Cabbages Plantation （Brassica oleracea var. capitata L.）

Subculture Helicoverpa armigera nuclear polyhedrosis virus （HaNPV） is an entomopathogenic virus isolated from cadaver of Helicoverpa armigera larvae, and is successfully produced in Spodoptera litura larvae as an alternate host. These viral agents have been used to control the population of three species insect pests Spodoptera litura, Crocidolomia pavonana and Plutella xyllostella, which were infested in cabbages plantation. The concentration of virus of 4 × 10^7 polyhedra/mL was sprayed in cabbage leaves 24 h before infested of larva and every four days after. The research used randomized design which consisted of one factor （three species of second instar larvae, S. litura, C. pavonana, P. xyllostella） and eight replications. The result showed that the three species of larvae were sensitive against subculture HaNPV infection. The mortality of each larval species （S. litura, C. pavonana and P. xyllostella） was 100%, 97.5% and 98.7%, respectively. Subculture HaNPV can be utilized to control the population of three species of insect pests in cabbage plantation.

Youth’s Attitude Formation in Youth Popular Music1 Subculture in China

This study was intended to analyze the youth’s attitude formation in youth popular music subculture of youth in China.One of the most universal and mysterious human activities is music.In Western industrialized nations,systematic instruction in harmony,counterpoint,rhythmic structure,composition,improvisation,and ear training is used to enhance the skill of musicianship.This course provides professional musicians with a variety of useful abilities that they may apply in their day-to-day work.There are various disciplines that are necessary to understand how music impacts human life,from cultural studies to political economics to sociology to psychology to acoustics.This study will use descriptive regressional method as its design and methodology.Descriptive research describes the phenomenon,condition,and situation.Various genres of youth popular music were found to be preferred by Chinese youth in their everyday lives,as well as the link between their idols and their musical education and their views about learning youth popular music in school.

Contemporary Youth Subcultures： The CDA Approach

In recent years, the cultural studies began to take a linguistic turn. This paper employed the three-dimensioned model of Fairclough, a critical discourse analysis （CDA） approach to explore youth subcultures. The cultural discourse of youth in different periods was examined in terms of discursive strategies, discursive practices, and social practices. The analysis shows that the subcultural groups adopt various discursive strategies to resist mainstream ideologies and construct the subcultural ideology and their collective identity. Meanwhile, their discursive practices build effective interaction mode within the subcultural groups. The examples of subcultures reveal that subcultures often emerge in the time of the social and cultural change and reshape the mainstream ideologies and unreasonable social structure.

Discursive Strategies of Contemporary Subcultures in the Global Context

Since the 20th century, the subculture, once a marginal cultural phenomenon, has been transformed into a global post-subculture. In this process, subcultural groups have undergone profound changes in multiple aspects. Hybridity, openness, and dissolution of depth become the keywords to understand youth subcultures. This paper is attempted to explore its discursive changes in comptemporary subcultures. By describing and interpreting the most popular subcultural discourse--hip-hop discourse, it is found that counter language often used in the subcultural discourse has been weakened and new strategies, such as code switching and semiotic games arise. The paper argues that with the transition of discursive strategies, subcultural groups attempt to weave the network of the global subcultures and create an interactive discursive space for global youth.

The Anime Subculture and Socialist Core Value System

The rise of the anirne subculture assembles the huge youth groups in the construction of the socialist core value system, and provides a new carrier and injected into the innovative dynamic. At the same time, their perverse behavior and exaggerated style of dress and the strange language constitute the alienation and challenges of a core value system. It should strengthen the ideological review and classification management, with the criticism of the mainstream media to enhance the vigilance of the community, as well as through commercial activities to digest rebel style, which is the effective way to promote the core value system.

Study on Tissue Culture of Leucophyta brownii ‘Canal Rocks Form’

[Objective] This study aimed to establish a tissue culture technology for Leucophyta brownii ‘Canal Rocks Form’. [Method] Leucophyta brownii ‘Canal Rocks Form’ stems with axillary buds were selected as explants in vitro to investigate the effects of different concentrations of 6-BA and NAA on proliferation culture and the effects of IBA, NAA and subculture cycle on rooting culture. [Result] The results showed that Leucophyta brownii ‘Canal Rocks Form’ plantlets need low concentrations of phytohormones and the rooting culture was significantly affected by the subculture cycle; the optimal medium for proliferation culture was MS＋0.5 mg/L 6-BA＋0.05 mg/L NAA＋30 g/L sucrose＋5.5 g/L agar, with a proliferation coefficient of 4.25; the plantlets with a subculture cycle of 28 d were the most suitable for rooting culture, with a rooting rate of 95.9% in 1/2 MS＋0.1 mg/L IBA＋0.05 mg/L NAA＋20 g/L sucrose＋5.5 g/L agar, and the average number and length of roots were 4.69 and 1.68 cm, respectively. [Conclusion] This study laid the foundation for establishing sterile culture system of Leucophyta brownii ‘Canal Rocks Form’.

Study on Tissue Culture and Rapid Propagation of Lycium ruthenicum Murr.

[Objective] This study aimed to establish a tissue culture and rapid propagation system for Lycium ruthenicum Murr.[Method] Using tender stems of L.ruthenicum as explants,MS as basic culture medium,the effects of different factors on primary culture,subculture and rooting of L.ruthenicum plantlets were investigated.[Result] The most appropriate medium for primary culture of L.ruthenicum was MS ＋ ZT 0.2 mg/L ＋ IBA 0.01 mg/L,in which axillary buds grew well and were rarely vitrified with the germination rate of 88.73%.In addition,ZT exerted significantly better effects on subculture and proliferation of L.ruthenicum plantlets than 6-BA.The most appropriate medium for subculture and proliferation of L.ruthenicum plantlets was MS ＋ ZT 0.15 mg/L ＋ IBA 0.01 mg/L,in which L.ruthenicum plantlets grew rapidly and robustly without vitrification,and the proliferation multiple reached 5.83 times.The most appropriate medium for rooting of L.ruthenicum plantlets was MS ＋ IBA 1.0 mg/L,in which the rooting rate reached 100%.The most appropriate substrate for transplanting and hardening of L.ruthenicum plantlets was humus soil：perlite = 1：1,in which L.ruthenicum plantlets grew well with the survival rate of 92.37%.[Conclusion] This study provided theoretical basis for largescale production and popularization of L.ruthenicum.

Optimization of rhizogenesis for in vitro shoot culture of Pinus massoniana Lamb.

The rooting capacity of Pinus massoniana is poor,especially for mature trees,and has prevented the development of clonal forestry for P.massoniana.In this study,we varied explant types,subculture times and exogenous hormones for plantlet regeneration and assessed shoots for rooting rate and root number for P.massoniana.Following fi ve repetitive grafts,new shoots from grafts used as explant sources were rejuvenated as observed from juvenile shoot morphology and anatomy,leading to greatly enhanced plant regeneration in comparison to that of mature materials from 26-year-old P.massoniana trees.The rooting capacity of subcultured shoots increased with successive subcultures,reaching a peak at 20 subcultures with 35–40 days per subculture.However,rooting performance was signifi cantly reduced after 30 subcultures.The addition of naphthaleneacetic acid(NAA)plus indoleacetic acid in the medium improved the root number,but the combination of exogenous NAA with paclobutrazol(PBZ)increased rooting rate and root number.We thus greatly improved the rooting capacity of mature P.massoniana trees by optimizing explant types(rejuvenated),subculture times(20 subcultures,35–40 days per subculture)and addition of NAA+PBZ to the rooting medium.The conditions can be used for effi cient plantlet regeneration of P.massoniana.

Nursing culture assessment tool (NCAT): Empirical validation for use in long-term care

Background:Capturing general aspects of the occupational subculture of nursing is needed in long-term care(LTC)given its latent influence on the quality of care that residents receive and on the ability of nursing staff(licensed nurses and certified nursing assistants)to implement evidence-based practice innovations.The psychometrically validated Nursing Culture Assessment Tool(NCAT)provides a comprehensive assessment using six dimensions(teamwork,communication,satisfaction,professional commitment,behaviors,and expectations),and evaluation of these dimensions could help positively reshape the culture before any change implementation.Purpose:Aims were to:(1)assess the validity and reliability of the NCAT across nursing staff in a single type of clinical setting e LTC facilities,and(2)present a refined theoretical model of the interaction of culture and practice implementation.Methods:A cross-sectional,exploratory investigation of the NCAT in LTC settings was conducted.Empirical construct validity of the 19-item NCAT's six subscales was investigated by confirmatory factor analysis using a sample of licensed nurses and certified nursing assistants(n?318).Results:The model fit was judged using the comparative fit index(0.94)and standardized root mean-square residual(0.05).Cronbach's alpha correlation coefficients of items in each subscale and in the overall scale ranged from 0.76 to 0.94.Conclusion:A summary of the NCAT development and report on its psychometric properties when administered in LTC settings is provided,extending previous findings of the NCAT's enhanced stability when used in assessing nursing staff perceptions in LTC and by demonstrating that the NCAT is a reliable and valid psychometric screening tool for nursing culture.

Study on Callus Induction from Immature Embryos and Plant Regeneration of Different Genotypes of Maize

The induction, subculture and differentiation of callus from immature embryos of maize （ Zea mays L. ） inbred line were studied and optimized. The re- suits revealed that, 2 mg/L 2,4-D was the optimal concentration to induce embryonic callus. Calli induced from inbred Qi319 and LY92 had good morphology and high regeneration. The embryos of the two inbred lines were selected as explants to establish efficient and stable tissue culture and transformation system.

Direct Organogenesis from Cotyledonary Node Explants of Cucurbita pepo (L.)—An Important Zucchini Type Vegetable Crop

Cotyledonary node explants from 3 - 5-d-old seedling derived explants of Cucurbita pepo (L.) produced multiple shoots in Murashige and Skoog (MS) salts B5 vitamins containing medium in the presence of N6-benzylamino-purine BAP 1 mg/L + Thidiazuron TDZ 0.05 mg/L. After 4 weeks explants were subcultured to medium containing MS salts B5 Vitamins BAP 0.5 mg/L, Gibberellic acid GA3 1 mg/L and L-Glutamine 15 mg/L. Periodic excision of regenerated shoots from the explants increased shoot efficiency during subculture. The combination of L-Glutamine with BA and GA3 significantly increased the shoot proliferation. Elongated shoots were excised from shoot clumps and transferred to rooting medium containing indole-3-butyric acid (IBA, 1.0 mg/L). The rooted plants were hardened in small pot containing standardized garden soil, well developed plant transferred to greenhouse and survival rate was 85%.

Cell Viability of Byrsonima intermedia A Juss Calli

Byrsonima intermedia A Juss. is a species with pharmacological properties from the Brazilian Cerrado that shows difficulties related to sexual propagation. Research on cell viability may provide useful information for the selection of cells with embryogenic potential during the callus culture, Within this context, our research is aimed at establishing the cell viability of calli from Byrsonima intermedia leaf segments. The calli went through three subculture phases, of 60 days each, in MS medium with 0.09 M sucrose, 0.6% agar, pH 5.8 and 4.52 laM 2,4-D. The calli were stored in dark conditions and samples were collected every 10 days from each subculture for viability tests with fluorescein 3,6-diacetate （FDA） and 2,3,5-triphenyltetrazolium chloride （TTC）. The staining methods allowed quantifying cell viability in each subculture. The best results from the FDA tests were obtained at 21, 25 and 29 days for the first, second and third subcultures respectively, with 53,86%, 61.88% and 53.73% viable cells. Regarding the TTC test, the largest absorbance values were obtained at 21, 27 and 28 days for the first, second and third subcultures respectively. Fluorescence and spectrophotometry analyses were efficient for determination of cell viability during callus cultivation period.

Research on the Multiple Exclusion Phenomenon in the Chinese“Core Weeaboo”Group

Among the subcultures that have developed rapidly in recent years,weeaboo culture covers a growing number of people.Meanwhile,corresponding differentiation has taken place among the weeaboos,typically including“core weeaboo”group and“pan weeaboo”group.Generally,“core weeaboos”refer to a group of people who are deeply involved in the weeaboo culture.On one hand,in the online community,“core weeaboos”tend to reject normal interaction with“pan weeaboos”,who they think are not deeply involved in the culture;on the other hand,due to the long-term prejudice of the mainstream culture against the weeaboos,the“core weeaboo”group has also suffered from the exclusion from the mainstream culture for a long time,thus forming a unique chain of“multiple exclusion”.With the“core weeaboo”group as the research object,this paper conducts online and field research on the phenomenon of“multiple exclusion”in the relevant groups by means of questionnaire and interview.It is found that the phenomenon of“multiple exclusion”in essence can be considered a two-way interaction between the“core weeaboo”group and the mainstream culture.In order to alleviate this phenomenon,this paper advocates that mainstream culture should increase its understanding of subcultural groups,and actively encourage high-quality“fusion”,so as to reduce the adverse effects on corresponding subcultural groups.

The Influence of Social Media on Contemporary Subcultural Groups in China： The Case of BL Fandom

Social media has become the infrastructure of contemporary virtual life and greatly influenced the landscapes of the modern culture. By tracking the development of a local subculture in China-BL fandom and analyzing BL fandom on social media, this research argues that social media empowers the subcultural groups and makes the cultural expression of BL fans heard. Meanwhile, the social relationship of online subcultural groups shows the tendency of decentralization and conglomeration, which is typical of social network services. In the interaction between fujoshies through social media, the subcultural groups also develop a collective identity resistant against the mainstream society. The rise of social media leads the contemporary subcultures to a new stage.