鲤鱼sGnRH基因克隆及其在成熟个体的表达分析

采用RACE方法,从鲤鱼脑组织克隆了两个差异的sGnRH(salmon GnRH[Trp^7Leu^8]GnRH)cDNAs,即cDNA1和cDNA2,其长度分别为393和478bp。两个cDNAs都包括一个285bp开放阅读框,编码的sGnRH前体为94个氨基酸残基,由一个信号肽、sGnRH十肽和一个由蛋白水解位点(Gly-Lys-Arg)连接的促性腺激素释放激素相关肽共3部分组成。用内含子捕获得到相应的两个差异sGnRH基因,即sGnRH genel和gene2,其基本结构都包括4个外显子和3个内含子,3个内含子的核苷酸相似性分别为71．1％、76．1％和88．0％。鲤鱼sGnRH cDNAs及基因的基本结构和编码特点与已报道的不同形式GnRH cDNAs和GnRH基因相似,由此推测所有类型的GnRH可能来自一个共同的祖分子。Southern杂交进一步证实鲤鱼基因组存在两个不同的sGnRH基因座位。相对定量RT-PCR检测发现,两个sGnRH基因除在精巢的表达存在差异外,在脑区、垂体和成熟卵巢共表达。其中两个sGnRH基因在端脑和下丘脑的表达水平明显高于后脑区。根据sGnRH mRNAs在多个脑区、性腺和垂体的共存推测,sGnRH可能对鲤鱼下丘脑-垂体-性腺轴的调节有至关重要作用,同时可能起神经调节剂或自分泌和旁分泌调节因子的作用。     

Sequences of cDNAs for human salivary and pancreatic α-amylases

The nucleotide sequences of the cloned human salivary and pancreatic α-amylase cDNAs correspond to the continuous mRNA sequences of 1768 and 1566 nucleotides, respectively. These include all of the amino acid coding regions. Salivary cDNA contains 200 bp in the 5′-noncoding region and 32 in the 3′-noncoding region. Pancreatic cDNA contains 3 and 27 bp of 5′- and 3′-noncoding regions, respectively. The nucleotide sequence humology of the two cDNAs is 96% in the coding region, and the predicted amino acid sequences are 94% homologous.Comparison of the sequences of human α-amylase cDNAs with those previously obtained for mouse α-amylase genes (Hagenbuchle et al., 1980; Schibler et al., 1982) showed the possibility of gene conversion between the two genes of human α-amylase.     

Structure of the dihydrofolate reductase gene in Chinese hamster ovary cells.

Overlapping recombinant lambda 1059 phages carrying regions of the dhfr locus from the amplified Chinese hamster ovary (CHO) cell clone MK42 have been isolated. In addition, dhfr cDNAs from this cell line have been cloned into plasmid pBR322. Restriction analysis of these recombinant molecules has led to a map of the Chinese hamster dhfr gene. This gene has a minimum size of 26 kb and contains six exons as defined by hybridization to a combination of mouse and CHO cDNA probes. The latter probes reveal 3' exonic sequences that are not present in mouse cDNA. The CHO dhfr gene thus extends about 700 bp further 3' than in the mouse, consistent with the larger size of the hamster mRNA. At least five intervening sequences are present, of approximate sizes: 0.3, 2.5, 8.6, 2.6 and 9.4 kb. Four sequences from highly repeated families are situated in introns within the dhfr gene. The overall structure of this gene is strikingly similar to that of the mouse. Evolutionary conservation of interrupted gene structure among mammals thus extends to genes that code for household enzymes as well as specialized or structural proteins.     

A novel promoter controls Cyp19a1 gene expression in mouse adipose tissue

Background  

Aromatase, the key enzyme in estrogen biosynthesis, is encoded by the Cyp19a1 gene. Thus far, 3 unique untranslated first exons associated with distinct promoters in the mouse Cyp19a1 gene have been described (brain, ovary, and testis-specific). It remains unknown whether aromatase is expressed in other mouse tissues via novel and tissue-specific promoters.     

Isolation and characterization of glucocorticoid- and cyclic AMP-induced genes in T lymphocytes.

Glucocorticoids and cyclic AMP exert dramatic effects on the proliferation and viability of murine T lymphocytes through unknown mechanisms. To identify gene products which might be involved in glucocorticoid-induced responses in lymphoid cells, we constructed a lambda cDNA library prepared from murine thymoma WEHI-7TG cells treated for 5 h with glucocorticoids and forskolin. The library was screened with a subtracted cDNA probe enriched for sequences induced by the two drugs, and cDNA clones representing 11 different inducible genes were isolated. The pattern of expression in BALB/c mouse tissues was examined for each cDNA clone. We have identified two clones that hybridized to mRNAs detected exclusively in the thymus. Other clones were identified that demonstrated tissue-specific gene expression in heart, brain, brain and thymus, or lymphoid tissue (spleen and thymus). The kinetics of induction by dexamethasone and forskolin were examined for each gene. The majority of the cDNA clones hybridized to mRNAs that were regulated by glucocorticoids and forskolin, two were regulated only by glucocorticoids, and three hybridized to mRNAs that required both drugs for induction. Inhibition of protein synthesis by cycloheximide resulted in the induction of all mRNAs that were inducible by glucocorticoids. Preliminary sequence analysis of four of the 11 cDNAs suggests that two cDNAs represent previously undescribed genes while two others correspond to the mouse VL30 retrovirus-like element and the mouse homolog of chondroitin sulfate proteoglycan core protein.     

The human insulin-like growth factor II gene contains two development-specific promoters

The insulin-like growth factors (IGF) play an important role in fetal and postnatal development. Recently, the nucleotide sequences of the cDNAs encoding IGF-I and IGF-II and part of the human IGF genes were reported. In this communication we describe two distinct IGF-II cDNAs isolated from a human adult liver and a human hepatoma cDNA library, respectively. Using these two cDNAs, we have established that the human IGF-II gene contains at least 7 exons. Two different IGF-II promoters have been identified, 19 kilobases (kb) apart, which are active in a development-specific manner. The promoter, active in the adult stage, is located only 1.4 kb downstream from the insulin gene.     

鳜鱼胰蛋白酶和淀粉酶与胃蛋白酶原基因的克隆与序列分析

采用RT-PCR及RACE法,克隆得到鳜鱼(Siniperca chuatsi)肝胰脏胰蛋白酶(trypsin, Try)、淀粉酶(amylase, Amy)基因 cDNA全序列.结果表明,鳜鱼Try基因cDNA全长为896 bp,其中开放阅读框 (open reading frame,ORF)为744 bp,编码247个氨基酸. 序列同源性分析发现,鳜鱼Try与 斑马鱼(Danio rerio)、非洲爪蟾(Xenopus laevis)、 小鼠Try和人TRY氨基酸序列同源性分别为81.4%、75.3%、74.5%和71.4%.鳜鱼Amy 基因cDNA全长为1 647 bp,其中ORF为1 539 bp,编码512个氨基酸.鳜鱼Amy与斑马鱼 、非洲爪蟾、小鼠Amy和人AMY氨基酸序列同源性分别为79.7%、75.4%、71.9%和70.9%. 同时对鳜鱼基因组进行PCR,获得鳜鱼Try、Amy与胃蛋白酶原(pepsinogen, Pep)全基因组DNA序列.序列分析表明,鳜鱼Try基因由4个内含子和5个外显子组成,全长1 362 bp;鳜鱼Amy基因由8个内含子和9个外显子组成,全长4 267 bp;鳜鱼Pep基因由8个内含子和9个外显子组成,全长 4 032 bp,与其它脊椎动物基因结构相似.应用Genome walker方法在鳜鱼克隆得到长度分别为1 189 bp、413 bp和527 bp的Try、Amy和Pep基因的5′侧翼区序列以及1段长为704 bp的Pep 基因3′侧翼区序列,并利用相关软件预测其中具有多个可调节其表达的调控元件.鳜鱼Try、Am y和Pep基因组全序列的克隆及其序列、结构分析和分子系统进化等的研究,为鱼类消化代谢相关基因的生理功能及表达调控机理进一步研究提供依据.     

Two Cyp19 (P450 aromatase) genes on duplicated zebrafish chromosomes are expressed in ovary or brain

Cytochrome P450 aromatase (Cyp19) is an enzyme catalyzing the synthesis of estrogens, thereby controlling various physiological functions of estrogens. We isolated two cyp19 cDNAs, termed cyp19a and cyp19b, respectively, from zebrafish. These genes are located in linkage groups 18 and 25, respectively. Detailed gene mapping indicated that zebrafish linkage groups 18 and 25 may have arisen from the same ancestral chromosome by a chromosome duplication event. Cyp19a is expressed mainly in the follicular cells lining the vitellogenic oocytes in the ovary during vitellogenesis. Cyp19b is expressed abundantly in the brain, at the hypothalamus and ventral telencephalon, extending to the olfactory bulbs. The expression of duplicated cyp19 genes at two different tissues highlights the evolutionary significance of maintaining two active genes on duplicated zebrafish chromosomes for specific functions in the ovary and the brain.