Two forms of cytochrome P450 cDNA from 3-methylcholanthrene-treated European eel Anguilla anguilla

ABSTRACT:     The cytochrome P450 (CYP) represents a large group of microsomal monooxygenases that catalyze drugs as well as a host of lethal environmental contaminants such as dioxins, leading to either detoxification and excretion from the animal or generation of carcinogenic intermediates. In the present study two forms of cDNA were cloned (Eu MC1 and Eu MC2) for European eel CYP1A genes by polymerase chain reaction (PCR) techniques. The cDNA of Eu MC1 was 3368 bp long coding 521 amino acid residues, and that of Eu MC2 was 2464 bp long coding 517 amino acid residues. Identities of deduced amino acid sequences between Eu MC1 and Japanese eel CYP1A1 and that between Eu MC2 and the second form of Japanese eel CYP1A were 98% and 97%, respectively, showing decisively that Eu MC1 and Eu MC2 are orthologous to Japanese eel CYP1A1 and the second form of CYP1A, respectively. A striking difference between the two eel species was that the Eu MC1 peptide was two amino acid residues longer than that of the Japanese eel CYP1A1. Existence of two loci of CYP1A in Japanese and European eels may suggest that the two forms of CYP1A exist widely among the eel species, because the divergence between the two eel species has been shown to be close to the basal divergence among eels. The identities in CYP1A may help to estimate genetic distance between European and Japanese eels.     

甜瓜果实中乙烯生产量及一些相关性状的遗传分析

为改进甜瓜货架期,利用成熟果实乙烯释放量及变化速率存在显著差异的3个甜瓜变种为亲本,配制了皇后×9-8(非跃变类型材料间)和皇后×Charentais(非跃变类型与跃变类型材料间)两个组合,对各组合F1,F2及回交世代BC果实中乙烯的生成变化及果实脱落性、外果皮迅速黄化等相关性状的遗传变化进行了分析。结果表明,甜瓜果实中乙烯生成量为不完全显性遗传,受两对以上基因控制。在皇后×Charentais组合的F1中,内源乙烯含量在采收1d后表现出急剧升高,同时伴随着外果皮迅速黄化及果柄脱落的发生,根据这些性状在F2及分别与父母本回交世代中的不同分离比率指出,乙烯的跃变类型、外果皮迅速黄化及果柄脱落在甜瓜果实中为显性遗传。     

水稻耐冷性研究Ⅲ.特定颖花结实率作为耐冷性指标的分子依据

以云南稻种耐冷性资源昆明小白谷及弱耐冷性的日本品种十和田配制的杂交F2代为材料,采用159个RFLP及SSR分子标记构建的连锁图和STATISTIC等分析软件,以单株结实率作为耐冷性评价指标.分析结果显示单株结实率与单株特定颖花结实率之间的相关系数(r)为0.8364;与耐冷性(以单株结实率为指标)相关的分子标记有43个,分布在8条染色体     

Characterization of Murine Pituitary-Derived Cell Lines Tpit/F1, Tpit/E and TtT/GF

The pituitary is an important endocrine tissue of the vertebrate that produces and secretes many hormones. Accumulating data suggest that several types of cells compose the pituitary, and there is growing interest in elucidating the origin of these cell types and their roles in pituitary organogenesis. Therein, the histogenous cell line is an extremely valuable experimental tool for investigating the function of derived tissue. In this study, we compared gene expression profiles by microarray analysis and real-time PCR for murine pituitary tumor-derived non-hormone-producing cell lines TtT/GF, Tpit/F1 and Tpit/E. Several genes are characteristically expressed in each cell line: Abcg2, Nestin, Prrx1, Prrx2, CD34, Eng, Cspg4 (Ng2), S100β and nNos in TtT/GF; Cxcl12, Raldh1, Msx1 and Twist1 in Tpit/F1; and Cxadr, Sox9, Cdh1, EpCAM and Krt8 in Tpit/E. Ultimately, we came to the following conclusions: TtT/GF cells show the most differentiated state, and may have some properties of the pituitary vascular endothelial cell and/or pericyte. Tpit/F1 cells show the epithelial and mesenchymal phenotypes with stemness still in a transiting state. Tpit/E cells have a phenotype of epithelial cells and are the most immature cells in the progression of differentiation or in the initial endothelial-mesenchymal transition (EMT). Thus, these three cell lines must be useful model cell lines for investigating pituitary stem/progenitor cells as well as organogenesis.     

Partial cDNA sequence of a relaxin‐like factor (RLF) receptor,LGR8 and possible existence of the RLF ligand‐receptor system in goat testes

Relaxin‐like factor (RLF), also known as insulin‐like factor 3 (INSL3), is produced by testicular Leydig cells, but its specific receptor LGR8 (leucine‐rich repeat family of G‐protein‐coupled receptor 8) has not been identified in goats. This study aimed to identify complementary DNA (cDNA) sequences of goat LGR8, and characterize the expression of both RLF and LGR8 in goat testes by RT‐PCR and immunohistochemistry. Testes were collected from immature (3‐month‐old) and mature (24‐month‐old) Saanen goats, and partial cDNA sequences of the goat homologue of human LGR8 were identified. The sequence encoded a reduced peptide sequence of 167 amino acids, which corresponded to transmembrane regions 2 through 5, followed by the beginning of intracellular loop 3 of human LGR8. Expression of both LGR8 and RLF genes was drastically increased in mature testes compared with immature ones. Although RLF protein was restricted to Leydig cells, LGR8 protein was detected in both Leydig cells and seminiferous epithelial cells (possibly germ cells and Sertoli cells). These results reveal a possible existence of the RLF‐LGR8 ligand‐receptor system within the goat testis, suggesting that RLF may play a role in testicular function through LGR8 on Leydig cells and seminiferous epithelial cells in an autocrine and/or paracrine manner.     

Isoenzyme polymorphism and segregation in isolates of Phytophthora infestans from Japan

Isoenzyme variation of 198 isolates of Phytophthora infestans collected from many locations in Japan during 1987–90, and of four pre-1987 isolates, was examined using starch gel electrophoresis. A previously unreported allele at malic isoenzyme locus/ME (90) was observed. An association between mating types and isoenzyme genotypes at three isoenzyme loci, glucosephosphate isomerase (GPI-1), peptidase (PEP-1) and ME, was found. At PEP-1, the A2 isolates from Japan had a previously unreported genotype (96/96). Normal segregation at the malic isoenzyme locus occurred in a cross of Japanese and Mexican parents. The results provide evidence of a change in the population genetic structure of P. infestans in Japan.     

The evaluation of the curd forming ability of milk replacers

Inadequate milk curd formation in the abomasum of newborn calves causes malnutrition and diarrhea. In order to define the factors of inadequate curd formation, we compared the curd forming ability among 9 kinds of milk replacers, bulk milk (raw milk), and skim milk both in vitro and in vivo . When rennet was added, the raw milk and one milk replacer formed firm curds. The rest of the milk replacers and skim milk did not form any curd. When a solution of HCl was added, raw milk, three milk replacers and skim milk formed the curd at pH 4.5, but the other milk replacers did not. When HCl was added following the rennet, raw milk, one milk replacer and skim milk formed the curd. In vivo , raw milk, two milk replacers and skim milk showed good curd formation whereas the other milk replacers showed poor curd formation inside the abomasums of the calves. This study showed that most of the milk replacers sold in Japan could not form the curd with rennet.     

Evaluation of the PATHFAST Chemiluminescent Enzyme Immunoassay for Measuring Progesterone in Whole Blood and Serum of Mares

Evaluation of a new chemiluminescent enzyme immunoassay, the PATHFAST assay system (PATHFAST), for measurement of circulating progesterone in mares was performed. Five mares at the mid-luteal stage were administrated a single i.m. injection of prostaglandin F2α analog (PGF2α; cloprostenol 250 μg/ml), and then blood samples were collected from the jugular vein at 0, 15, 30 and 45 min, at one-hour intervals until 24 and at 48 hr via a catheter in the jugular vein. To monitor the physiological changes in circulating progesterone in mares after induced luteolysis, concentrations of progesterone in whole blood and serum samples were measured by PATHFAST. In addition, concentrations of progesterone in serum samples measured by PATHFAST were compared with those measured by radioimmunoassay (RIA) and enzyme immunoassay (EIA). Using PATHFAST, the serum concentrations of progesterone in mares correlated highly with those of whole blood samples (r=0.9672, n=88). The serum concentrations of progesterone as measured by PATHFAST correlated well with RIA (r=0.9654, n=88) and EIA (r=0.9323, n=112). An abrupt decline in circulating progesterone in whole blood samples was observed within 2 hr (50%), followed by a gradual decline until 48 hr later. The results for progesterone in whole blood samples correlated highly with those in serum samples, and the declining pattern paralleled that of the serum samples. These results demonstrated that PATHFAST is useful in the equine clinic as an accurate diagnostic tool for rapid assay of progesterone within 26 min, using unextracted whole blood.