Lithophyllum okamurae is one of the important encrusting coralline algae, which plays important roles as primary producer, carbonate sediment builder, and habitat provider in the marine ecosystems. In this study, L. okamurae was collected from tropical coast of Sanya, and firstly described based on both detailed morph-anatomical characteristics and molecular studies of typic DNA sequences. The structure of the thalli of L. okamurae was pseudoparenchymatous construction with radially organized dimerous organizations in the crustose portion. The pseudoparenchymatous construction were composed of three parts, including 1 to 3 layers of epithelia cells which had flatten to round outermost walls, one layer of square or rectangular cells of the hypothallia and multiple layers of square or elongated rectangular peripheral cells. Palisade cells were observed, and the cells of the contiguous vegetative filaments were connected by secondary pit-connections with cell fusions absent. The carposporangial conceptacles, the spermatangial conceptacles, the bisporangial conceptacles and the tetrasporangial conceptacles were observed, and all these four kinds of conceptacles were uniporate. The spermatangial conceptacles were slightly convex and buried at shallow depths in the thalli tissues, and the carposporangial conceptacles and asexual conceptacles were protruding and conical. Phylogenetic studies based on DNA barcoding markers of 18S rDNA, COI, rbcL and psbA revealed that L. okamurae clustered with the closest relation of L. atlanticum, and formed a distinct branch. Based on the comparative anatomical features and the molecular data, the detailed description of the valid species of L. okamurae was firstly given in this study to provide theoretical basis for algae resources utilization and conservation in marine ecosystems.

鲳属鱼类相似的外部形态特征导致其种类鉴定非常混乱，其中镰鲳模式标本的准确信息和DNA条形码更是无法获得。2010年6月到2013年4月期间，我们在丹东、东营、青岛、南通、舟山、温州、长乐、台湾及和歌山县（日本）收集了207尾镰鲳样品。基于上述样品归纳了镰鲳的主要形态鉴别特征为：背鳍Ⅷ-Ⅺ-43~51；臀鳍Ⅴ-Ⅷ-43~49；胸鳍22~27；尾鳍19~22，腹鳍缺失。第一鳃耙细弱，稀疏，3~4+12~16=15-20。脊椎骨数为39~41。头部后上方侧线管的横枕管丛和背分支丛后缘呈弧形，略短，背部横枕管丛末端未达胸鳍上部基点。腹部横枕管丛稀少，明显短于背部横枕管丛。通过比较NCBI上下载的COI同源序列，两个显著的组群被检测到，研究结果显示在氨基酸序列和组群间遗传距离的基础上，该两大组群实为两个有效种，其中组群1的序列可以作为镰鲳的DNA条形码。本论文提出镰鲳正确的外部形态特征和DNA条形码，可为后续鲳属鱼类的研究提供高效、准确的保障，也为其分类提供理论基础，并能鲳属鱼类的渔业管理和保护策略提供合理的参考。

Codium, one of the largest marine green algal genera, is difficult to delimit species boundary accurately based on morphological identification only. DNA barcoding is a powerful tool for discriminating species of seaweeds. The plastid elongation factor TU (tufA) is considered as maker to perform DNA barcoding of green algal species than rbcL gene due to universality and rapid evolution rate. We conducted DNA barcoding application to Codium specimens from the Jeju Island, Korea to overcome the limit of morphological identification and to confirm the species diversity. As a result of applying tufA marker, we newly generated fifty-five tufA barcodes to resolve eight species. TufA marker exhibited 6.1%-21.8% interspecific divergences, wider than the gap of rbcL exon 1, 3.5%-11.5%. Molecular analysis of rbcL exon 1 sequences of Codium revealed eight distinct species like tufA analysis separated in five phylogenetic groups. DNA barcoding of the genus Codium using tufA marker is more helpful to overcome the limit of morphological identification, and this is more potential to reveal cryptic species and to resolve the relationships among subspecies than rbcL analysis alone. The complement of tufA barcoding and rbcL analyses including morphology for the genus Codium in the northwestern Pacific will give much more reliable achievement for discovering species diversity and resolving the phylogenetic relationships.

Identification of hydrozoan species is challenging, even for taxonomic experts, due to the scarcity of distinct morphological characters and phenotypic plasticity. DNA barcoding provides an efficient method for species identification, however, the choice between mitochondrial cytochrome c oxidase subunit Ⅰ (COI) and large subunit ribosomal RNA gene (16S) as a standard barcode for hydrozoans is subject to debate. Herein, we directly compared the barcode potential of COI and 16S in hydrozoans using 339 sequences from 47 pelagic hydrozoan species. Analysis of Kimura 2-parameter genetic distances (K2P) documented the mean intraspecific/interspecific variation for COI and 16S to be 0.004/0.204 and 0.003/0.223, respectively. An obvious "barcoding gap" was detected for all species in both markers and all individuals of a species clustered together in both the COI and 16S trees. These results suggested that the species within the studied taxa can be efficiently and accurately identified by COI and 16S. Furthermore, our results confirmed that 16S was a better phylogenetic marker for hydrozoans at the genus level, and in some cases at the family level. Considering the resolution and effectiveness for barcoding and phylogenetic analyses of Hydrozoa, we strongly recommend 16S as the standard barcode for hydrozoans.

In contrast to typical planktonic hydromedusae, Cladonema medusae are mostly benthic, with specialised adhesive branches to adhere to the substrate. In this study, a Cladonema species discovered in a laboratory aquarium in Fuzhou, China, was confirmed as a new species, based on morphological and molecular analyses. The species was named Cladonema multiramosum sp. nov. Its medusa is distinct from that of congeners possessing substantially more adhesive branches (8–24, rarely 5–7), and tiny branches on the upper radial canals. The validity of C. multiramosumum sp. nov. was also supported by molecular phylogenetic analyses based on the mitochondrial 16S rDNA sequence. However, C. multiramosum sp. nov. medusa also displayed considerable phenotypic plasticity with respect to its radial canals, tentacles, stinging branches per tentacle, oral tentacles, manubrium, and gonads, hindering species identification based solely on morphology. Although some morphological characteristics of hydroids (filiform tentacles and medusa buds) and nematocysts could also be used as diagnostic characters in the genus Cladonema, this information is unavailable for some Cladonema species. Thus, the taxonomy within the genus Cladonema was re-evaluated based mainly on the morphological characteristics of the medusa. Further revision of the genus Cladonema should be made when supplementary information on the life cycle and DNA barcoding are updated.

Two morphotypes of Plectropomus leopardus have been identified; morphometric and meristic analyses show that there is no diagnostic difference between them. A difference in color pattern was the most appropriate phenotypic character with which to distinguish between the two morphotypes. Complete mitochondrial DNA sequencing, however, indicated a clear difference between the two morphotypes. Barcoding analysis revealed no significant difference (P>0.05) in CO1 or ND2 divergence among intramorphotypic individuals, even between geographically separated populations, whereas the intermorphotypic CO1 and ND2 divergences were large enough (averaging 0.95% for CO1 and 1.37% for ND2) to clearly discriminate between the two morphotypes. The color pattern difference, geographical distribution, together with the mtDNA and barcode sequencing data, suggest that the two morphotypes should be of two subspecies or even two species.

Somatic cells were prepared from sea snail enzyme digests of Porphyra yezoensis thalli.Using SDS-Proteinase K as extraction solution,total DNA was isolated from the somatic cells.The crude extracts of total DNA were purified with glassmilk,and the resulting DNA was of sufficient quality for digestion of restriction endonuclease.DNA bands were clearly observed in the restriction patterns of EcoRI,PstI and HaeⅢ respectively.The presence of DNA bands in the restriction pattern of total DNA indicated that the genome of Porphyra yezoensis may be small.Unexpectedly,using guanidinium isothiocyanate and sarcosyl as extraction solution,a plasmid-like DNA band(2.3 Kb) was directly found in the isolated total DNA of Porphyra yezoensis.A very simple and convenient method for plasmid-like DNA isolation has been established.

RAPD (Randomly Amplified Polymorphic DNA) analysis was performed with filaments of 15 Porphyra lines representing four important groups (P.yezoensis,P.haitanensis,P.katadai var.Hemiphylla and P.oligospermatangia:Eight stable and repeatable RAPD bands amplified with two primers,OPN-02 and OPJ-18,were selected for the construction of DNA fingerprinting.The RAPD results were scored based on the presence or absence of each of the 8 bands and then converted to computer language expressed with two digitals,1 and 0,which represented the presence (numbered as 1) or absence (numbered as 0) of each band,respectively.Based on these results,a model DNA fingerprint and a computerized DNA fingerprint were constructed.In the constructed DNA fingerprint,each Porphyra line has its unique fingerprinting pattern and can be easily distinguished from each other.Later,a software,named as PhGI,was designed based on this DNA fingerprinting.It can be used in practical Porphyra line identification.

This article describes in detail the procedures in obtaining spermatids from living animals and in isolating sperm nuclei DNA by hydroxyapatite chromatography.The isolated DNA is free of RNAs and proteins after treatment with proteinase K, followed by chromatography on hydroxyapatite column which is eluted with stepwise-increased concentrations of potassium phosphate buffer.More than 89% of the DNA purified is found in the eluates of both 0.2 M and 0.25 M potassium phosphate buffer.The two fractions are identical in ultraviolet absorption spectrum.DNA molecules thus prepared should be useful for studying DNA structure and DNA replication by electron microscopy, since the length of the purified DNA molecules is in the range of the size of a replicating unit.

A protocol to coextract the microbial metagenomic DNA and RNA from deep-sea sediment was developed for the microbiological study of environmental samples.The obtained pure metagenomic DNA with the size larger than 23 kb and stable RNA could be used directly for PCR and reverse transcription-PCR(RT-PCR)respectively.The direct lysis including the treatments of SDS,proteinase and lysozyme was applied to acquiring the metagenomic DNA and RNA furthest.Prior to the lysis treatment,the glass bead and denaturing solution were added to enhance the lysis efficiency and keep the integrity of RNA respectively.Denaturing gradient gel electrophoresis(DGGE)was applied in accessing the microbial 16S rRNA diversity by PCR and RT-PCR amplification from a single extraction.The pattern obtained by this analysis revealed some differences between them,indicating the efficiency of the protocol in extracting the metagenomic DNA and total RNA from deep-sea sediment.