Application of RAPD in genetic diversity study onGracilaria lemaneiformis III. Phase and sex related markers

Random amplification of polymorphic DNA (RAPD) was conducted by using 10 random primers (P-1 to P-10) inGracilaria lemaneiformis. Phase and sex specific bands were amplified by primers P-2, P-6, P-7 and P-8: for P-2 a 1.4 kb band was found in female gametophytes and tetrasporophytes, for P-6, a 0.6 kb band appeared in male gametophytes and tetrasporophytes; for P-7, a 0.76 kb band appeared in male gametophytes and tetrasporophytes; for P-7, a 0.72 kb band appeared in female gametophytes and tetrasporophytes; for P-8, a 0.73 kb band only appeared in male gametophytes.

     

RAPD study on some common species ofPorphyra in China

Electroporation, PEC, PEG plus electroporation and Biolistics methods were tested in gene transformation ofP. yezoensis. The exogenousgus was from plasmid of pBI121 and pCAMBIA1301, both contain the CaMV35S promoter. The receptors included the protoplasts, tissues and free-living conchocelis filaments ofP. yezoensis. Several factors, for example, the voltage, capacitance and bivalent cations, etc., were studied. Results show that these four methods are all efficient for gene transformation inP. yezoensis; and that PEG is the best one, with transformation efficiency of up to 4×10⁻⁵. GUS activity was detected 26 days after transformation by using PEG method.

     

Comparative studies on the larval development of the penaeid shrimps,Penaeus Chinensis,P. merguiensis andP. penicillatus

The morphology of larval and 1st postlarval stages ofPenaeus penicillatus are described. Results from comparative studies on larval development ofP. penicillatus, P. merguiensis andP. chinensis are as follows: These three species could not be identified during their naupliar stages. In the 1st protozoea, the antennule L1/L2 value is 1.7–2.0 inP. merguiensis, but less than 1.7 inP. chinensis andP. penicillatus; in the 2nd protozoea, the supra-orbital spine inP. chinensis is not bifurcated, while those ofP. merguiensis andP. penicillatus are bifurcated; in the 3rd protozoea, there is a minute (or no) dorso-median spine on the posterior margin of the 1st and 2nd abdominal somite inP. chinensis, but they are prominent inP. merguiensis andP. penicillatus. In the mysis and 1st postlarval stages,P. chinensis differs fromP. merguiensis andP. penicillatus in having 9 (8 in the other 2 species) long setae on the exopod of pereopods 1–3; additionally, one dorsal tooth appears on the rostrum ofP. chinensis in the 2nd mysis and that of the other 2 species in the 3rd mysis;P. chinensis has 2 (mostly) or 1 dorsal tooth on the rostrum in the 3rd mysis and 2–3 in the 1st postlarva, whileP. penicillatus andP. merguiensis have only 1 in the 3rd mysis and 1st postlarva. Comparative studies on larval development showedP. penicillatus has closer affinity withP. merguiensis than withP. chinensis. Contribution No. 2218 from the Institute of Oceanology, Chinese Academy of Sciences     

Study on monospore-like cells fromPorphyra haitanensis

Porphyra haitanensis andP. yezoensis are two mainPorphyra species cultured in China. Their life histories are slightly different. So far we have not observed thatP. haitanensis naturally produces monospores developing into thalli.P. yezoensis produces monospores which directly germinate into young thalli used in cultivation (Zeng, et al., 1985). Some somatic cells inP. yezoensis develop in vitro into monospore-like cells which later grow into young thalli (Lu, 1983). Studies on whether or not somatic cells inP. haitanensis can produce in vitro monospore-like cells that later grow into young thalli is important for understanding its life history and for culturing new varieties.     

Iodine content ofSargassum in Southern China

Sodium thiosulphate and neutron activation determination of iodine content in samples of 18Sargassum species collected in Guangdong and Guangxi Province in April 1996 showed large differences (0.47×10⁻³ inSargassum hemiphyllum, 0.56×10⁻³ inSargassum assimile, and 4.5×10⁻³ inSargassum vachellianum). Traditional titration analysis and neutron activation analysis for determination of total iodine yielded similar results showing that certainSargassum species had high capacity to accumulate iodine. Contribution No. 3162 from the Institute of Oceanology, Chinese Academy of Sciences. The project was supported by the National Ninth Five-Year Plan of China (No. 96-916-04-01).     

Construction of a new gene integration platform system for theSynechococcus sp. PCC7942

According to the known sequence of iron stress-induced gene (isiAB operon), we cloned its 1.5 kb fragment by PCR, and used this fragment as integration homologous fragment. After several steps of subcloning donor DNA into theisiAB fragment, a donor plasmid pZL which could be integrated into the chromosomal DNA ofSynechococcus sp. PCC7942 was constructed. In order to express the heterologous gene at a high level through the integration platform system, we constructed the donor DNA by the following steps. We cloned the strong promoter (240 bp) of heat shock genegroESL operon fromSynechococcus sp. PCC7942 by PCR. Then subcloned the multiple cloning sites (MCS),rbcS polyA into the downstream of thegroESL promoter. The kanamycin resistance gene, as the marker gene, was also subcloned into the donor DNA. Thus, in the donor plasmid pZL, the integration homologous fragment and several expression elements, such asgroESL promoter, MCS,rbcS polyA terminator and kanamycin resistance gene, were all included.

After naturally transformed and introduced the donor plasmid pZL intoSynechococcus sp. PCC7942, as in the pZL, the donor DNA sequence is flanked by two DNA fragments (0.4 kb and 0.7 kb) homologous to theisiAB fragment ofSynechococcus sp. PCC7942, the homologous DNA can recombine with the chromosomal DNA. After screening by kanamycin, the transformants which integrated the heterologous DNA were selected. The efficiency of transformation is about 1×10⁻⁶. By southern blot analysis, it was confirmed that the donor DNA had been integrated into the chromosomal DNA ofSynechococcus sp. PCC7942, located on the site of theisiAB gene, and can be replicated with the chromosomal DNA.

     

The genetic diversity,individual relatedness and possible mating system of an isolated population of the Cyprinid species Megalobrama pellgrini in upper reaches of the Changjiang (Yangtze) River,China

Megalobrama pellegrini is a cyprinid fish endemic to upper reaches of the Changjiang(Yangtze) River, China, which is also an important economic species in the local area. In recent years,resources of this species have decreased sharply and its conservation has drawn great attention. In the present study, we collected 120 individuals from the Longxi River, a tributary isolated from the main channel of the Changjiang River, where M. pellegrini is still relatively abundant. Using two different molecular markers, mitochondrial cytochrome b(cyt b) gene, and nuclear microsatellite(simple sequence repeat,SSR), we analyzed the genetic diversity of this isolated population. The results show that sequence genetic diversity was low(H_d:0.290 and P_i: 0.000 77 for cyt b gene),while the SSR genetic diversity was high(H_o:0.824 4±0.147 2,H_e: 0.823 510.145 1). Analysis indicated that this population had experienced a bottleneck,with inbreeding and small effective population size(around 50). Based on SSR data, relatedness analyzing revealed that the 120 samples were grouped into 10 completely independent clusters. It was inferred that the mating system of M. pellegrini was polygamy. We suggested that the low genetic diversity could be induced by the overfishing and inbreeding depression. Therefore, we suggested that the urgent conservation measures should be taken to control the overfishing and give better conditions for the fish to grow and spawn, then to restore population size.     

Morphological changes in development of pharyngeal teeth inMylopharyngodon piceus

Adult pharyngeal teeth inMylopharyngodon piceus are molariform. Based on SEM observations of the developing teeth, this paper describes the morphological diversification of pharyngeal teeth inM. piceus. The larval and juvenile teeth are changed from conical to adult molariform teeth through seven stages. Comparisons are made between each stage and corresponding types in some species of different subfamilies in Cyprinidae. It is considered that the ontogenetic resemblances of the pharyngeal teeth bear relationship to the phylogeny of cyprinids. This project was funded by the International Cooperation of Japan—China and the National Natural Science Foundation of China     

NEW OBSERVATIONS ON THE MEIOTIC PROCESS IN THE MARINE DINOFLAGELLATE NOCTILUCA SCINTILIANS （NOCTILUCALES, DINOPHYCEAE）

The meiotic process in Noctiluca scintillans were observed under light microscope.Some abnormal cell divisions, incompletely separated “zoospores” and the changes of the zoospores are described in this paper. Together with the fmdings of field samplings and the previous results by other researchers, the process of meiosis in N. scintillans was supposed to be a pathway to reduce the extra high density of NH3-N within the cell in order to ensure normal population growth.     

CYTOLOGICAL COMPARISON OF GAMETOGENESIS OF SCALLOPS, ARGOPECTEN IRRADIANS AND CHLAMYS FARRERI

Histological characteristics of gametogenesis of two kinds of scallops, gonochoric Chinese scallop, Chlamys farreri and hermaphroditic bay scallop, Argopecten irradians were investigated in this study. Spermatogenesis in C. farreri has different developmental stages: spermatogonia, primary spermatocyte, second spermatocyte, spermatid and spermatozoon. A large number of same developmental stage spermatic cells converge at a definite area of the testis. Premeiotic, previtellogenic and vitellogenic oocytes can be found during oogenesis in C. farreri, where oocyte distribution is obviously irregular. The A. irradians gonad consists of two different parts in one individual: one part functions as testis, the other as ovary. Between these two parts is a special appearance area, where a large number of spermatic cells are bound with two layers of acellular substance with many oocytes in it.     

Averaged sound intensity in shallow water having a negative leap-layer and a constant depth

The leap-layer is assumed to consist of two isovelocity water layers. The expressions of the law on the decay of the averaged sound intensity with range are derived when the source and receiver are laid in the same isovelocity water layer or in two different isovelocity water layers respectively. The results show that there is apparent depth constitution in the sound field in the leap-layer. The theoretical results agree quite well with the practical data. This paper was published in Chinese inOceanologia et Limnologia Sinica 15(6): 550–557, 1984.     

A transformation model forLaminaria Japonica (Phaeophyta,Laminariales)

A genetic transformation model for the seaweedLaminaria japonica mainly includes the following aspects:

1. 1.

   The method to introduce foreign genes into the kelp,L. japonica

   Biolistic bombardment has been proved to be an effective method to bombard foreign DNA through cell walls into intact cells of both sporophytes and gametophytes. The expression ofcat andlacZ was detected in regenerated sporophytes, which suggests that this method could induce random integration of foreign genes.

   Promoters to drive gene expression

2. 2.

   The CaMV35S promoter was first used by us to induce the expression of GUS gene in brown algae. But results of further studies suggested that CaMV35S could be a tissue-specific promoter. Our use of SV40 promoter resulted in both transient and stable expression oflacZ andcat in sporophytes or gametophytes. No GUS or LacZ background was found in either sporophytes or gametophytes.The regeneration route of transgenic kelp

   The regeneration efficiency of explants is still very low. By using female gametophytes as gene hosts and parthenogenesis as regeneration route, CAT activity and LacZ activity were detected in regenerated sporophytes of parthenogenetic kelp. li]4.|The way to select transgenic kelp

3. 1.

   Results of sensitivity tests showed that kelp was only sensitive to chloramphenicol and hygromycin among many antibiotics. The regenerated sporophytes by parthenogenesis were more sensitive to hygromycin than to chloramphenicol. Resistant kelp was created by transforming female gametophytes with pSV40-CAT and stimulating parthenogenesis followed by selection in medium with lethal concentration of chloramphenicol.

   Safety consideration of transgenic kelp

   L. japonica was originally introduced from Japan. In China it is a cultured population. The possibility of its negative impact on natural populations is very low. 2) The vectors and target genes used for transformation should be restricted in order to avoid any negative impacts on human health and environment. 3) Specially devised containers (3.6 L, made of 200 μm membrane) were used to ensure that the kelp cannot escape or be eaten by marine animals. 4) To avoid the release of spores, it is very necessary to harvest the kelp at suitable age before the sporangium forms.

     

Analysis of biological geochemistry of chemical elements in Betula ermanii forest in Changbai mountains, China

Based on catalogue of biology and geochemistry of chemical elements, content characteristics and variation law of the large nutrient elements, the needful trace elements, the uncertain needful elements, the non-needful elements and the toxic elements in Betula ermanii trees are analyzed. The result shows that the content of the large nutrient elements in Betula ermanii trees is higher than that of other kinds of element; the contents of all kinds of elements in foliage with vigorous metabolism are higher than those in other parts; the content variations of the large nutrient elements and the needful trace elements with similar chemical property, geochemical property and biological function in different parts of Betula ermanii trees show the similar laws; but the other three kinds of elements variations are without laws. It is indicated that the variation of the needful elements in the plant follows a certain law, they are in relative equilibrium under undisturbed condition. Foundation item: Under the auspices of the Natural Science Foundation of the Chinese Academy of Sciences. Biography: LIU Jing-shuang(1956 -), male, a native of Jilin Province, professor, doctor tutor. His research interest includes environment science. E-mail: liujingshuang@mail.ccig.ac.cn     

Effects of Mg2+, NaCl, citric acid, and other factors on synthesis and accumulation of β-carotene inDunaliella salina

The effect of Mg²⁺, NaCl and citric acid on the accumulation of β-carotene inDunaliella Salina was studied. The experimental results showed that 10.5 mmol/L Mg²⁺, 5 mol/L NaCl, 3 μmol/L citric acid, and CO₂ are favorable forDunaliella Salina cell growth and β-carotene accumulation. After 144 h culture under the above conditions, theDunaliella Salina biomass increased by 7.18 times; β-carotene reached 9.61%. Project 89023990 supported by the Shandong Natural Science Fund.     

The spectrum characteristics of Bohai Bay wind waves under the influence of cold waves

The wind waves generated by cold waves are some of the most dangerous waves in Bohai Bay. In this paper records of the wind waves under the influence of cold waves in November, 1968, are discussed. It is shown that the north wind is very strong in spring and winter. The frequency spectrum of the development and decline of wind waves depends mainly on the wind speed. The change of shape and scale parameters of the spectrum are discussed and the calculation equations are shown. This paper was published in Chinese inOceanologia et Limnologia Sinica.16(2): 147–155, 1985.     

The relation of the average sound intensity to range in homogeneous shallow-water

Using a “three parameters” bottom model, the average sound intensity in homogeneous shallow water with constant depth is found by a better expression to be finite for normal mode number of water layer. The space structure of the sound field and some factors controlling the near, middle and far fields are discussed with this expression. This paper was published in Chinese inOceanologia et Limnologia Sinica 14 (2): 198–202, 1983.     

Cell biotechnology of rhodophytes in China

Cell biotechnology of rhodophytes is important not only in theoretical research but also in cultural practice and for exploitation of genetic resources. In this paper, cell biotechnology ofPorphyra is reviewed. Tissue culture and protoplast studies on other rhodophytes in China are discussed.

     

Climatic Signals Recorded in Shells in Xingcuo Lake,Eastern Tibetan Platedu

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Influences of wind on vertical structures of temperature and salinity,and upwelling off Zhejiang coast in summer

In this paper the influences of short-term wind on vertical structures of temperature and salinity, and the upwelling off the Zhejiang coast are studied on the basis of hydrographic data and wind near the coast in Summer (1959–1979). It is pointed out that the variations of vertical distributions of temperature and salinity, and upwelling in this region are closely related to short-term (several days to a week) variation of wind observed. The occurrence of double thermocline, a special oceanographic phenomenon, is partially related to unstable SSW wind field. SSW and NE winds in June are two dynamic factors making intermediate warm water form or disappear. Contribution No. 792 from the Institute of Oceanology, Academia Sinica. This paper was published in Chinese inOceanologia et Limnologia Sinica 14 (1): 14–21, 1983.     

Cloning and sequencing of the ferredoxin gene of blue-green algaAnabaena siamensis

The structure gene for ferredoxin,petFI, fromAnabaena siamensis has been amplified by polymerase chain reaction(PCR) and cloned into cloning vector pGEM-3zf(+). The nucleotide sequence ofpetFI has been determined with silver staining sequencing method. There is 96.8% homology between coding region ofpetFI fromA. siamensis and that ofpetFI fromA. sp. 7120. Amino acid sequences of seven strains of blue-green algae are compared.