2009 Vol. 30, No. 5
Actins are a small family of ubiquitous proteins that are essential cytoskeletal components and are highly conserved during evolution. Actins are usually divided into two classes, the cytoplasmic and muscle actins, which have different functional roles. Here we systematically analyzed the actin genes in the genome of the primitive chordate amphioxus (Branchiostoma floridae). We found that amphioxus contains more than 30 actin genes, many of which are linked. Phylogenetic analysis suggests the amphioxus actin genes have clearly undergone extensive expansion through tandem duplications. The actin genes’ structure also varies a lot, containing 2 to 7 exons. We also cloned two muscle type of actin genes from the amphioxus（B. belcheri）and compared their expression patterns during early development. The slight difference in their expression suggests functional diversification of these actin genes. Our results shed light on the evolution both of actin genes themselves and their functional roles in chordate development.
Leptin, the product of the obese gene, is one of the key factors for the regulation of food intake and whole-body energy balance. Both grass carp (Ctenopharyngodon idellus) and silver carp (Hypophthalmichthys molitrix) leptin full-length cDNA were cloned by RT-PCR and RACE methods. The sequences of leptin cDNA in grass carp and silver carp were 1 096 bp, 1 176 bp, and the deduced protein had 173, 172 amino acids respectively. The amino acid homologues analysis showed high identity in cyprinid fish. Despite lower amino acid identity between cyprinid fish and other leptins, conservation of both cysteine residues that formed leptin’s single disulfide bridge inside the chain presented a certain similarity. Phylogenetic tree showed that leptin in grass carp and silver carp were in the same branch as other fishes. The 5′-flanking region and introns of leptin in grass carp and silver carp were characterized using Genome Walker and PCR methods. The genes were 2 129 bp and 2 192 bp in length respectively in grass carp and silver carp, with similar gene structures in other vertebrates (including 3 exons and 2 introns). This research will be helpful for further studies of fish leptin in cultivation and food safety.
2009, 30(5): 487-496. doi: 10.3724/SP.J.1141.2009.05487
The sequences of the control region (D-loop) of mitochondrial DNA ( mtDNA) were obtained from five species of sturgeon. The sequences obtained had lengths between 795-813 bp. The motifs of the conserved sequence block (CBS) and termination-associated sequence (TAS) were identified in the sequences. Examination of the molecular phylogeny using maximum likelihood, maximum parsimony and Bayesian analysis shows that sturgeons form two clades representing biogeographic distribution, and the Huso is not monophyletic. Heteroplasmy in 150 fishes was due to a variable number of tandem repeats. Length of the central repeat units ranges from 78 to 82 bp with repeat sequence copies between four and six. The repeat units, followed by an imperfect repeat unit, are in high similarity between species, with 82.93% between Amur sturgeon and starlet and 90.59% between Siberian sturgeon and Russian sturgeon. Furthermore, comparing with other repeat sequences published, different species have the same repeat unit and intramolecular recombination event may happen in the repeat unit. All of these indicate that the repeat units are related to the evolution of sturgeon and arose prior to species divergence after which recombination happened.
The epithelial cell of the Mongolian sheep's fallopian tubes were cultured as in vitro model, different concentrations of progesterone (10-6, 10-7, 10-8, 10-9 and 10-10 mol/L) were added in the culture, and then the relative expressions of sheep β-defensin-1 (sBD-1) mRNA were surveyed by real time PCR. The results showed that, compared with the control group, the relative expressions of sBD-1 mRNA added with 10-6 and 10-7 mol/L progesterone were significantly higher (P<0.05), and those of 10-8 and 10-9 mol/L groups were remarkably higher (P<0.01). However, there was no significant difference between 10-10 mol/L group and control. Among groups added with progesterone, the sBD-1 mRNA expressions of 10-8 and 10-9 mol/L groups were markedly higher than that of 10-10 mol/L group (P<0.01), but there was no difference among other added groups. The results indicated that progesterone promoted the relative expression of sBD-1 mRNA in the epithelial cell of the fallopian tubes within the indicated concentrations (10-9-10-6 mol/L). And the relative expression of sBD-1 mRNA was varied with the different concentrations of progesterone. In conclusion, the expression of β-defensin was related to progesterone in female reproductive tract.
2009, 30(5): 503-508. doi: 10.3724/SP.J.1141.2009.05503
Calpains are the key proteases involved in muscle growth, protein transformation and meat tenderization. In this study, CAPN7 cDNA of Wild boar was cloned by RT-PCR, its sequence analysis, expression characterization and polymorphism analysis were performed. The results showed that the full-length coding sequence of Wild boar CAPN7 is 2442 bp long, and encodes a protein of 813 amino acids with the catalytic site of calpain family, whose catalytic triad residues is located at amino acids 290, 458 and 478, respectively. RT-PCR expression analysis indicated that CAPN7 mRNA was ubiquitously expressed in the 12 tissues examined, but the relative abundance was unevenly distributed in tissues. The CAPN7 mRNA levels in muscle of 6 and 9-month-old crossbred pigs were higher than those of Yorkshire pigs at the same ages. In addition, a missense mutation was identified by PCR-SSCP and its distribution in wild boar, Min pig and Duroc was significantly different. The results lay the foundation for further studying the function of CAPN7 gene.
Embryonic stem cells (ESCs) hold great potential for the treatment of liver diseases. Here, we reported the induction of rhesus monkey ESCs along a hepatocyte lineage by a three-step method. The Rhesus monkey ESCs were suspended in DMEM medium supplemented with 10% FBS for 11 days to form embryonic bodies (EB) which contained early endoderm or hepatic cells by expression of early hepatic genes. EB of 11 days were transferred into ECM coated dishes and cultured with aFGF, BMP-4 and OSM at different time points for further studies. After induction by aFGF and BMP-4 for 7 - 10 days, the differentiated cells showed a hepatocyte-like morphology, expressed hepatic marker genes (such as AFP，ALB APOH，G-6-P and TAT) and proteins (AFP，ALB and CK18), and possessed the capacity to storage intracytoplasmic glycogen. After further induction with OSM for 7 - 10 days, some cells shared similar characteristics of matured hepatocytes, such as expression of genes of CYP1B1 and ADH1C, and ICG uptaken, indicating the presence of functional hepatocytes in the differentiated cells.
Previous studies have confirmed that both honeybee and Drosophila are capable of learning and memory. This study aimed to investigate whether the house fly (Aldrichina grahami), with strong instincts to adapt their living environment, have the learning ability to associate odor stimulus to avoid electric shock in free flying state using a device developed by the authors. The result showed the learning ability of Aldrichina grahami at the electric shock voltages of 5 V, 25 V and 45 V AC. When 60 V was used, the flies were frequently injured. Our results indicate that Aldrichina grahami is a good model to study the neural mechanism of learning and memory. The paradigm in this study has some advantages that can be used in future studies of free insects.
In this report, we have developed efficient methods for total RNA extraction from hepatocarcinoma tissues and cell lines, reverse transcription,cDNA cloning and quantitive real-time PCR (q-RT-PCR). We compared the effect of two reverse transcriptase (M-MLV, SuperScriptII) on the efficiency of total RNA reverse transcription and four DNA polymerase(Taq polymerase，Pfu polymerase, LA taq polymerase，Prime Star polymerase) in cloning large fragment of cDNA. The impact of different RNA integrity on q-RT-PCR and large fragment of cDNA cloning had also been investigated in this study. The newly developed RNA extraction method greatly improved the integrity of total RNA. We found that the integrity of total RNA is vital to large fragment cDNA cloning. RNA degraded at some extent is only suitable for q-RT-PCR analysis but not for cDNA cloning.
Using the open-field and Y-maze behavioral paradigms to assess the novelty-seeking behavior of the male Sprague-Dawley rats that received bilateral electrolytic lesions of the orbitofrontal cortex (OFC) or sham lesions, we examined the role of the OFC on exploration of the novel environment. In the open-field task, the rats with OFC lesions exhibited the reduced average ambulation distance and average rearing number when compared with the animals with sham lesions. In the meanwhile, the rats with OFC lesions showed the less duration of visits and number of entries in the novel arm in the Y-maze task than the control animals. The current findings suggest that the OFC plays an important role on the novelty-seeking behavior in rats.
Significance of plasticity in energy budget and development of animals in response to variations of food availability was examined in weaned male KM mice that were acclimated to a random food deprivation (FD) for 4 weeks, and then refed ad libtum for another 4 weeks. Food intake was determined using a food balance method. Basal metabolic rate (BMR) was measured using a closed-circuit respirometer. Food intake increased significantly on ad libitum day in FD mice, but BMR and activity decreased. FD mice also had a significantly lower weight in carcass and gonadal gland than controls after 4 weeks of FD. All the above parameters recovered to the levels of controls after 4 weeks of ad libitum refeeding, indicating a significant plasticity. In addition, the group difference in fat content was not significant. These results suggest that animals can compensate for unpredictable lower food availability by an energetic strategy, including an increase in food intake and a decrease in energy expenditure associated with BMR and activity, and decrease in carcass mass thus reduced energy spent on maintenance, but not including the changes in body fat. Development is affected significantly by lower food availability but recovers to the normal level when food is plentiful. Finally, plasticity in energetic budget and development play important roles in animals under unpredictable variations of food availability.
Through confirming the feeding characteristics and daily feeding amount of Hoolock gibbon (Hoolock hoolock), we discussed whether daily feeding amounts are different among individuals and seasons. Factors impacting daily feeding amount and characteristics were analyzed. According to results stated above, a base will be established for studying daily energy requirements and nutritional carrying capacity of Hoolock gibbon. The gibbons were observed and data were recorded by focal animal sampling method at Nankang of Mt. Gaoligong in spring and autumn from 2007 to 2008. The results showed that gibbons ate 40 parts from 36 botanic species. The feeding characteristics about food unit, feeding speed and weight of one food unit were different between fruits and young leaves. In spring and autumn, the percentage of fruits in daily feeding amount was the highest, the percentage of leaves was second, and the percentage of stems and flowers was the lowest. The daily feeding amount of the female in the family group was more than that of the male in spring and autumn. The daily feeding amount of the female solitary gibbon in spring was slightly more than that of the female in the family group. In spring, a feeding bout for feeding fruits by the solitary gibbon was twice longer than that of the family group, and the time for feeding young leaves was almost equal between the solitary gibbon and the family group. The daily feeding amount of the same individual in autumn was more than in spring, and the percentage of different parts of plants in daily feeding amount was also different between in spring and in autumn. The results showed that diet and amount of daily feeding part reflected the basic strategy for food selection. The size of individual between male and female, energy costs, food characteristics and requirements of physiology were factors impacting the daily feeding amount of Hoolock gibbon. The feeding bout of the gibbon was interrelated with amount of food resource in feeding tree.
Relative fatness is an important general indicator of physiological body condition in small mammals and reflects their adaptive ability to the environment. In May 2008, we surveyed part islands in Qiandao Lake about the number of populations of small mammals through the night trapping method, anatomizing and measuring the dominant species Niviventer confucianus captured during the search, calculating the index of relative fatness K, (K=100W/L3), where L indicated the body length, and W was the carcass body weight; Meanwhile we investigates the relationship between the relative fatness and the changing characteristics in sex, age, habitat types and other factors under islanding conditions. Results indicated that islanding affected the Niviventer confucianus fatness between the sexes and the extent of their differences ranged from significant level (large islands, P=0.015) to no significance (small and medium islands, P=0.063 and 0.288); among age groups, only a few adjacent age groups had significant difference (large: ♂，I and II, P=0.042；media：♀and ♂ all II and III, P=0.013 and 0.044) or very significantly (large: ♀, III and IV, P=0.000), the others failed to meet the significant level (P>0.05). The changes of Niviventer confucianus fatness among age groups were mostly related to the changes of its own physiological characteristics; Isolated island resulted in significant difference in different types of dominant species and abundance, thus affecting the difference of fatness; There was significant difference (P=0.000) between different sizes islands and had significant positive correlation between them (Pearson test，r=0.561，P=0.037). In a word, in addition to sex and age, the islands area, habitat and population pressure all have a direct or indirect relationship with the difference of fatness in Qiandao Lake, which also reflects the adaptability of Niviventer confucianus to islanding environment.
In order to understand the foraging behavior and ecology of Chinese crocodile lizard (Shinisaurus crocodilurus), we studied its prey discrimination mechanisms through two series experiments. In the first experiment, the swabs were dampened by deionised water, cologne, smashed earthworm (Pheretima sp.) and smashed tenebrio (Tenebriomolitor L.) as the chemical cues and observed the response of 11 Chinese crocodile lizard. Each individual was tested 24 times to each stimulus.The results showed that all individuals of Chinese crocodile lizard responded to the swabs by tongue flicking and the number of tongue-flicks in response to tenebrio and earthworm stimuli were significantly higher than that to cologne and deionized water (Wilcoxon test, all P<0.001). It indicate that the Chinese crocodile lizard can discriminate food and non-food stimulus. In the second experiment, we observed the responses of Chinese crocodile lizard to the following cues: (A) blank utensil, (B) the utensil was wetted by smashed earthworm, (C) the alive earthworm was sealed in the utensil, (D) alive earthworm was kept in the open utensil. Each individual was tested 5 times in each cue. Results showed that the Chinese crocodile lizard for each cue was significantly different in dealing time, investigation frequency and attack frequency(all P<0.001). The dealing time, investigation frequency and attack frequency in cue C（only visual cue）and in cue D （visual and chemical cue）were significantly high than in cue B（only chemical cue）（all P<0.001). Without visual cue, the dealing time and investigation frequency were significantly high than in cue A (blank)(all P<0.001). Under the same visual cues, only dealing time in cue D was higher than that in cue D (Z=3.95, P<0.001), but investigation frequency and attack frequency were not different between cue C and D (for former Z=1.53, P=0.13；for latter Z=1.10, P=0.27) . These indicate that Chinese crocodile lizard rely more on visual cues to discriminate prey than on chemical cues. The prey discrimination mechanisms of Chinese crocodile lizard may be related with foraging mode, phylogeny and prey.
In this study, we experimentally examined the ovipositing sequence of the pollinator Ceratosolen fuscicep, non-pollinating wasp Platyneura mayri of Ficus racemosa and non-pollinating wasp Acophila sp.1 and Wakerella benjamini of Ficus benjamina in Xishuangbanna from Apr. 2006 to Jun. 2007. For the pollinating fig wasps, we injected ether into the receptive fruits with different time intervals after introducing the fig wasps, killing the fig wasps; for the non-pollinating fig wasps, we manually controlled ovipositing time length through organdy bags. After the treated fruits developed to be mature stage, we collected the adult wasps, and then analyzed and compared the offspring sex ratio under different ovipositing time length. The data showed that fig wasp C. fuscicep, P. mayri and Acophila sp.1 firstly oviposit their male offspring. The male offspring was much higher than female, if the mother wasps only oviposited one hour, whist the female offspring will be increased with the increase of oviposting time of mother wasps and the female offspring will be much higher than male offspring when the mother wasps could sufficiently oviposit their offspring in these three fig wasps. These results partly explain that the female offspring sex ratio decrease with increase of number of foundresses. However, the ovipositing sequence of Wakerella benjamini randomly oviposit their eggs and the sex ratio is about 1:1. Our results here imply that the ovipositing sequence and ovipositing time length of mother wasps might be of the most important factors determining the offspring sex ratio of fig wasps.
The nest-site selection of White-crowned Penduline Tit Remiz coronatus in Northern Xinjiang was studied from April to July in 2008. It has special nesting behavior. The nest has a pocket pouch size, with a delicate structure. Research of the nest, using the method of total area investigation, included searching for nests carefully, combined with tag plotting method, and drawing the nest distribution map. In total 125 nests were found in the field, nesting on willow, poplar, birch and other hardwood trees near lakes and rivers. 68.80% of nest trees were willows Salix spp. The average height of nests was 5.3±2.5 m. The nests were located in the lower part of arbor (about 1/3), about 70% of nests less than 30 meters from river. Study of the nest-site selection, used a quadrate survey to investigate the parameters of nest-site characteristics. The principal component analysis indicated that there were four factors affecting the nest-site selection of the White-crowned Penduline Tit. They were: 1. canopy (including diameter of nest tree and canopy cover above nest), 2. nest tree species (including species of nest tree and arbor, height of nest tree and nest in the tree), 3. position or site (including distance to river and nest orientation ), 4. food and nest material (including canopy cover under nest).
The community characteristics of soil Sarcomastigophora in the Mayan Forest Region of the National Nature Reserve of Xiaolong Mountains was studied using the “non-flooded petri dish method” and observation in vivo from July of 2006 to April of 2007. In total 78 species were identified, including 2 unnamed species and 12 new records of soil Sarcomastigophora in China, belonging to two subphylums，five classes, 16 orders, 32 families, and 47 genera. Amoebida was the dominant group, Kinetoplastida and Euglenida were the subdominant groups, Dinoflagellida, Chaenoflagellida, Pelobiontida, Diplomonaida and Actinophyrida were the incidental group, and Bodo minimus, Bodo ovatus, Oikomonas termo and Raphidiophrys viridis were the dominant species. The result of the study showed that there were abundant species and endemic and rare species in the community of soil Sarcomastigophora, the species composition of soil Sarcomastigophora in the Mayan Forest Region of the National Nature Reserve of Xiaolong Mountains was unique. The community characteristics of soil Sarcomastigophora was discussed based on analyzing the complexity and uniqueness of the ecological environment in the Mayan Forest Region of the National Nature Reserve of Xiaolong Mountains.
Bar-headed geese (Anser indicus) is one of important migratory goose’s species group in Qinghai lake region, the population dynamics of which is the foundation data to decision-making of Bar-headed geese’s protection in Qinghai lake National Nature Reserve in future. A bar-headed goose population dynamics model was constructed in Qinghai lake region regarding the dynamics with STELLA graphical modeling software. The parameters included in the model that describing mating, egg deposition, hatching, brood-rearing, immigrating, emigrating and death of bar-headed goose had been set on the base of long time studied and observed data of bar-headed goose. In running simulation models, we use one year as unit and 1/7 as a time step in accordance of periodic time of bar-headed goose stay in Qinghai lake .The next 25 years’ simulated population changes results of bar-headed goose showed that trend of the maximum bar-headed goose population will vary around 20 thousand in Qinghai lake region. Meanwhile, the population was simulated that assuming calamity strikes in 2010.The model and simulated results about bar-headed goose population are reliable according calibrated results using observed data from 2006 to 2008, and the method of modeling bar-headed goose population provides with some reference to further research on other birds in Qinghai lake region. In the end, some suggestions of protecting bar-headed goose from danger have been put forward based on above results in paper.
In Nov 2006 and Oct 2008, members of the Kunming Institute of Zoology (KIZ) took surveys of fishes in the Niulan River, a tributary of Jinsha River, Yunnan, and collected 42 fish species. According to the historical record and the fish specimens preserved in the collection room of KIZ, there are a total of 59 fish species in Niulan River, which belong to 46 genera, 12 families and 5 orders. Ichthyologic fauna of Niulan River were separately analyzed by traditional sorting method and the Value of Faunal Presence (VFP) method at the respective levels of order, family and genus. Two kinds of different ranking methods get two rather different results. Traditional sorting analysis indicated that some world-distributed families and genera had a small proportion in the Niulan River. On the contrary, the VFP of some endemic families and genera turned out to be relatively high. The families Amblycipitidae and Channidae have two genera and which distribute in Africa and south Asia, with the highest VFP. The genera Jinshaia and Metahomaloptera, which distribute in the Yangtze River, have a VFP of 100%. Pseudogyrinocheilus and Oreias have one species, with a VFP of 100%. The VFP method can reflect better the character of ichthyologic fauna than the traditional sorting method. By combination of these two methods, we analyzed the ichthyologic fauna of Niulan River, and then we compared it with the lower reaches of Jinsha River ichthyologic fauna from Shigu to Yibin, which indicated high comparability.