malM, a new gene of the maltose regulon in Escherichia coli K12. I. malM is the last gene of the malK-lamB operon and encodes a periplasmic protein

The structure and expression of the distal part of the malK-lamB operon in Escherichia coli was studied. DNA sequencing was performed as far as a HinfI restriction site located 1313 base-pairs downstream from gene lamB. The open reading frame, formerly called molA, which begins 245 base-pairs downstream from gene lamB, is longer than was initially thought, and was renamed malM. It could encode a protein of 306 amino acid residues. The complete malM open reading frame was cloned under control of the tac 12 promoter. In maxicells, the resulting plasmid permitted tac12-promoted synthesis of two polypeptides, encoded by gene malM, with apparent molecular weights of 37 X 10(3) and 34.5 X 10(3). We provide strong evidence that the 34.5 X 10(3) Mr protein is derived from the 37 X 10(3) Mr protein by processing at the amino-terminal end, and that this processed form is located in the periplasmic space. We show that the chromosomal malM gene is expressed as part of the malK-lamB operon, and that its product is periplasmic. Finally, we demonstrate with nuclease S1 mapping experiments that the mRNA terminates at a typical rho-independent terminator located about 45 base-pairs beyond the end of gene malM, which is thus the last gene of the malK-lamB operon.     

Simultaneous expression of salivary and pancreatic amylase genes in cultured mouse hepatoma cells.

The tissue-specific expression of two types of mouse amylase genes does not overlap in vivo; the Amy-1 locus is transcribed in the parotid gland and the liver, while expression of Amy-2 is limited to the pancreas. We identified a mouse hepatoma cell line, Hepa 1-6, in which both amylase genes can be simultaneously expressed. Amy-1 is constitutively active in these cells and is inducible by dexamethasone at the level of mRNA. We demonstrated that the liver-specific promoter of Amy-1 is utilized by the dexamethasone-treated hepatoma cells, and that glucocorticoid consensus sequences are present upstream of this promoter. Amy-2 is not detectable constitutively, but can be activated if the cells are cultured in serum-free medium containing dexamethasone. Expression of Amy-2 in a nonpancreatic cell type has not previously been observed. We speculate that induction of Amy-1 and activation of Amy-2 may involve different regulatory mechanisms. Hepa 1-6 cells provide an experimental system for molecular analysis of these events.     

Interstrain Variation in Amylase Gene Copy Number and mRNA Abundance in Three Mouse Tissues

Amylase expression in strain YBR differs in several respects from the standard mouse phenotype. The synthesis of salivary amylase is elevated twofold in YBR mice and the synthesis of pancreatic amylase is reduced to one-half the normal rate. We have compared the concentrations of amylase mRNA in the parotid, liver and pancreas of YBR mice with those in strains A/J and C3H. We observed differences in amylase mRNA abundance which can account for the levels of amylase protein synthesis in the parotid and pancreas of these strains. Unexpectedly, the concentration of amylase mRNA in the liver of YBR mice was also higher than in the other strains. Since liver amylase is transcribed from the same gene as parotid amylase, duplication of the Amy-1 locus could account for the elevated mRNA concentration in both tissues. Quantitative analysis of genomic DNA by Southern blotting provided direct evidence for duplication of Amy-1 in strain YBR.     

Molecular analysis of the heterogeneity region of the human ribosomal spacer

The human ribosomal non-transcribed spacers are 30 X 10(3) base-pairs (or 30 kb) in length with a limited length heterogeneity localized in a specific region downstream from the 3' end of the transcribed region. Total DNA digested with EcoRI and BamHI and hybridized with a probe containing the 3' end of the 28 S ribosomal RNA coding region shows four major bands of 3.9 kb, 4.6 kb, 5.4 kb and 6.2 kb. The 5.4 kb band is the most abundant in every individual, followed by the 4.6 kb band. The longest and the shortest size classes are less well-represented and may even be absent. Every individual shows his own pattern of relative abundance of non-transcribed spacer length classes that can be followed through generations. We decided to investigate the molecular structure of the heterogeneity region, in order to cast light onto the mechanisms underlying the origin and maintenance of this length heterogeneity. Pertinent spacer regions of eight ribosomal clones from two human genomic libraries were subcloned and analyzed by restriction mapping and nucleotide sequencing. In the minimal length class, there is a sequence of 700 base-pairs that appears to be tandemly duplicated once, twice or three times in the other length classes. This repeated DNA module contains a region consisting of repetitions of simple pyrimidine groups like C-T, C-T-T-T or C-C-C-T. DNA module repeats may differ by the length of this pyrimidine-rich region. However, these length variations are not continuous, as revealed by Southern transfer analysis of several individuals and different cloned gene units: instead, the repeated modules fall into two discrete length classes of about 700 base-pairs and 800 base-pairs. An imperfect duplication of a short sequence of 86/89 base-pairs is present at the boundary between the heterogeneity region and the upstream flanking region, representing a very ancient duplication event.     

选育过程中的粤黄鸡血液淀粉酶（Amy—1）基因频率的世代变化

本文调查了粤黄鸡两个选育方向和交配方式均不同的品系的血液淀粉酶(Amy-1)各世代的分布情况,并以不同Amy-1类型的粤黄鸡作了交配试验。发现两个品系各世代都始终保持Amy-1分布不平衡状态,这种分布不平衡可能由各种Amy-1基因型生活力不同引起;另外,人工选择也可能造成Amy-1基因频率的改变。交配方式是否对Amy-1基因频率的改变起作用值得进一步研究。     

家蚕Wnt信号通路下游关键基因Pangolin转录剪接体X3的克隆和分析

【目的】克隆家蚕Bombyx mori Wnt信号通路下游关键基因Pangolin isoforms A/H/I/S转录剪接体X3 (Pangolin X3),分析其序列和表达特征。【方法】从NCBI数据库检索家蚕Pangolin X3,根据其编码序列(coding sequence, CDS)设计引物,利用PCR从家蚕幼虫中肠和血淋巴中进行克隆并测序验证。利用SilkDB 3.0, SMART,多序列比对和系统发育树分析Pangolin X3的序列特征。利用qRT-PCR分析Pangolin X3在家蚕5龄第3天幼虫不同组织(头、血淋巴、体壁、性腺、中肠、前部丝腺、中部丝腺、后部丝腺、脂肪体和马氏管)中的相对表达水平。【结果】从家蚕幼虫中肠和血淋巴克隆了Pangolin X3(GenBank登录号：XM＿038020921)的CDS,其开放阅读框长1 560 bp,编码519个氨基酸残基,预测分子量为55.86 kD,预测等电点为7.53。Pangolin X3蛋白含有保守的β-catenin结合位点和HMG结构域,其氨基酸序列在不同的昆虫中比较保守,特别是与DNA结合的HMG结构域...     

Molecular analysis of the Adh region of the genome of Drosophila melanogaster

A small region of the genome of Drosophila melanogaster has been cloned in a series of overlapping phage. A length of 165 X 10(3) base-pairs of contiguous DNA that spans polytene chromosome region 35A4 to 35B1 and includes the structural gene for alcohol dehydrogenase (Adh) as well as at least two other genes, outspread (osp) and no-ocelli (noc), has been characterized by mapping chromosome aberrations to the DNA. The relationship between osp and Adh is surprising: of nine osp alleles associated with chromosome breakpoints, five map distal (i.e. 5') to Adh and four map proximal (i.e. 3') to this gene. None affects the expression of Adh. As defined by these and other breakpoints, the osp gene spans at least 52 X 10(3) base-pairs and overlaps the Adh gene. The noc gene, as defined by the mapping of nearly 30 breakpoints, is at least 50 X 10(3) base-pairs in size. Alleles of noc and noc- deletions show either of two kinds of interaction with the recessive lethality of l(2)br29ScoR+1, a lethal that maps immediately distal to noc. One class of noc allele is viable when heterozygous with ScoR+1, while the other class is lethal or semi-lethal. Both classes, however, are homozygous or hemizygous viable. The locations of these two classes of noc allele on the DNA fall into two clusters, with those that are viable with ScoR+1 located proximal to those that are not. The physical boundary between these classes lies at a site just distal to that of the breakpoint of the inversion associated with ScoR+1 itself.     

A single mouse alpha-amylase gene specifies two different tissue-specific mRNAs

The alpha-amylase mRNAs which accumulate in two different tissues of the mouse, the salivary gland and the liver, are identical except for their 5' non-translated sequences: the 5' terminal 158 nucleotides of the major liver alpha-amylase mRNA are unrelated to the 5' terminal 47 nucleotides found in its salivary gland counterpart. DNA that specifies the 5'terminal one-quarter of these mRNAs has been isolated through genomic cloning and sequenced. The initial 161 nucleotides of the liver alpha-amylase mRNA are specified by DNA sequences that lie 4.5 kb upstream from those for the common body of the two mRNAs. In contrast, the 5' terminal 50 nucleotides of the salivary gland alpha-amylase mRNA are found 7.5 kb from sequences that the two mRNAs share in the genome. These cloned DNA sequences occur once per haploid genome, indicating that both the salivary gland and liver alpha-amylase mRNAs are transcribed from the same gene (Amy1A). Since no rearrangement of these DNA sequences can be detected among mouse sperm, salivary gland or liver preparations, gross rearrangement does not account for the tissue-specific pattern of expression observed for Amy1A. Rather, these data indicate that the salivary gland and liver alpha-amylase mRNAs are differentially transcribed and/or processed from identical DNA sequences in different tissues.