The 5′ untranslated region of Arabidopsis thaliana calmodulin cDNA is an independent cDNA containing an open reading frame

A cDNA clone, pAUK1, with an open reading frame (ORF) coding for a hypothetical 164-amino-acid protein was isolated from an Arabidopsis thaliana (L.) Heynh cDNA library. The clone was attached, tail to tail, to the 3′ end of A. thaliana hexokinase cDNA. An almost identical sequence had been previously described as the 5′ untranslated region (5′ UTR) of A. thaliana calmodulin cDNA (ACaM-2). Sequence comparison with three additional A. thaliana truncated cDNA clones which appear in a database (GenBank) supports the conclusion that pAUKl is identical to the 5′ UTR of ACaM-2 and that the 5′ UTR of ACaM-2 is an independent cDNA artificially linked to A. thaliana calmodulin cDNA.     

Characterisation of a pea hsp70 gene which is both developmentally and stress-regulated

A pea pod cDNA library was screened for sequences specific to lignifying tissue. A cDNA clone (pLP19) encoding the C-terminal region of a hsp70 heat shock protein hybridised only to pod mRNA from pea lines where pod lignification occurred. Expression of pLP19 was induced by heat shock in leaves, stems and roots of pea and chickpea plants. Four different poly(A) addition sites were observed in cDNAs derived from the same gene as pLP19. This gene was fully sequenced; unlike most hsp70 genes, it contains no introns. The 5-flanking sequence contains heat shock elements and other potential regulatory sequences.     

东亚飞蝗中肠几丁质酶基因的克隆、序列分析及组织定位

通过RACE方法,克隆了东亚飞蝗Locusta migratoria manilensis (Meyen)几丁质酶基因 (LmChi)cDNA全序列 (GenBank 登录号：EF092841)。获得的cDNA全长1 604 bp,其中可读框1 452 bp, 编码483个氨基酸。推测其氨基酸序列与18家族昆虫几丁质酶有较高的相似性。与其他几丁质酶一样,东亚飞蝗几丁质酶序列也包含一个信号肽、一个几丁质酶活性位点、一个碳端丝氨酸富集区和一个几丁质结合域。半定量RT-PCR研究表明,LmChi基因只在东亚飞蝗不同发育阶段的中肠组织中表达,而在东亚飞蝗体壁、前肠和后肠均没有发现LmChi基因的转录。     

Isolation and characterization of a pea pod cDNA encoding a putative blue copper protein correlated with lignin deposition

Nearly isogenic pea lines were selected to investigate the geneticbasis for lignification of the pea (Pisum sativum) pod endocarp.The development of the pod endocarp in the normal and mutantpea pod pheno-types was examined by light microscopy. A peapod cDNA library representing poly (A)+ RNA purified from L59pea pods (genotype, PV; phenotype, lignified endocarp) was differentiallyscreened with total cDNA probes prepared from total pod RNAfrom L59 and L1390 (genotype, pv; phenotype, no lignificationof endocarp) pods 4-6 d after flowering (DAF). A clone, designatedpLP18, was selected for further characterization on the basisof hybridization to the L59 cDNA probe, but not the L1390 cDNAprobe. Northern blotting was used to show that pLP18 representeda mRNA of 0.95 kb. The predicted polypeptide from the LP18 cDNAencoded a putative blue type 1 copper protein. The expressionpattern of LP18 mRNA in pods of the experimental pea lines wasdetermined using RT-PCR quantitation. Hybridization of the cDNAto pea genomic DNA showed that this protein is probably encodedby a single gene. Key words: Pisum sativum, pod endocarp, lignification, blue copper protein     

Transposon and spontaneous deletion mutants of plasmid-borne genes encoding polycyclic aromatic hydrocarbon degradation by a strain of Pseudomonas fluorescens

Pseudomonas fluorescens strain LP6a, isolated from petroleum condensate-contaminated soil, utilizes the polycyclic aromatic hydrocarbons (PAHs) naphthalene, phenanthrene, anthracene and 2-methylnaphthalene as sole carbon and energy sources. The isolate also co-metabolically transforms a suite of PAHs and heterocycles including fluorene, biphenyl, acenaphthene, 1-methylnaphthalene, indole, benzothiophene, dibenzothiophene and dibenzofuran, producing a variety of oxidized metabolites. A 63 kb plasmid (pLP6a) carries genes encoding enzymes necessary for the PAH-degrading phenotype of P. fluorescens LP6a. This plasmid hybridizes to the classical naphthalene degradative plasmids NAH7 and pWW60, but has different restriction endonuclease patterns. In contrast, plasmid pLP6a failed to hybridize to plasmids isolated from several phenanthrene-utilizing strains which cannot utilize naphthalene. Plasmid pLP6a exhibits reproducible spontaneous deletions of a 38 kb region containing the degradative genes. Two gene clusters corresponding to the archetypal naphthalene degradation upper and lower pathway operons, separated by a cryptic region of 18 kb, were defined by transposon mutagenesis. Gas chromatographic-mass spectrometric analysis of metabolites accumulated by selected transposon mutants indicates that the degradative enzymes encoded by genes on pLP6a have a broad substrate specificity permitting the oxidation of a suite of polycyclic aromatic and heterocyclic substrates.     

Cloning and expression pattern of a hemolin homologue from the diamondback moth, Plutella xylostella

Hemolin has been known to play a key role in insect innate immunity. In an attempt to examine expression pattern of the Hemolin gene in the diamondback moth, Plutellea xylostella, the full-length cDNA of Hemolin was cloned using 5′-RACE PCR technique. The cDNA contained a 5′ untranslated region of 48 nucleotides and a 3′ untranslated region of 198 nucleotides, including a stop codon (TAA) and a poly (A) tail. It consists of 1,401 bp with an open reading frame of 1,245 bp, encoding 414 amino acids. The deduced amino acid sequence of PxHemolin has relatively low identities (35?42%) to various insect Hemolins. However, it has high three-dimensional structural similarity to Hemolin. Interestingly, analysis of spatial expression pattern of PxHemolin shows that it was highly expressed in the Malpighian tubule and the silk gland although it was also detected in fat body and gut. Furthermore, PxHemolin mRNA was highly induced 3 hr after immune-challenging with lipopolysaccharide and was gradually up-regulated after laminarin treatment. These data suggest that PxHemolin may play a role in innate immune responses although it remains to further elucidate the precise biological functions in P. xylostella.     

The photoconvertible water-soluble chlorophyll-binding protein of Chenopodium album is a member of DUF538, a superfamily that distributes in Embryophyta

Various plants possess hydrophilic chlorophyll (Chl) proteins known as water-soluble Chl-binding proteins (WSCPs). WSCPs exist in two forms: Class I and Class II, of which Class I alone exhibits unique photoconvertibility. Although numerous genes encoding Class II WSCPs have been identified and the molecular properties of their recombinant proteins have been well characterized, no Class I WSCP gene has been identified to date. In this study, we cloned the cDNA and a gene encoding the Class I WSCP of Chenopodium album (CaWSCP). Sequence analyses revealed that CaWSCP comprises a single exon corresponding to 585 bp of an open reading frame encoding 195 amino acid residues. The CaWSCP protein sequence possesses a signature of DUF538, a protein superfamily of unknown function found almost exclusively in Embryophyta. The recombinant CaWSCP was expressed in Escherichia coli as a hexa-histidine fusion protein (CaWSCP-His) that removes Chls from the thylakoid. Under visible light illumination, the reconstituted CaWSCP-His was successfully photoconverted into a different pigment with an absorption spectrum identical to that of native CaWSCP. Interestingly, while CaWSCP-His could bind both Chl a and Chl b, photoconversion occurred only in CaWSCP-His reconstituted with Chl a.