Molecular cloning and characterization of a LEAFY-like gene highly expressed in developing soybean seeds

2007 ◽  
Vol 17 (4) ◽  
pp. 297-302 ◽  
Author(s):  
Qingchang Meng ◽  
Chunhong Zhang ◽  
Fang Huang ◽  
Junyi Gai ◽  
Deyue Yu
Keyword(s):  
Transcription Factors ◽  
Seed Development ◽  
Genomic Structure ◽  
Soybean Seed ◽  
Reproductive Organs ◽  
Pcr Analysis ◽  
Amino Acid Residues ◽  
Reading Frame ◽  
Rapid Amplification ◽  
Onion Epidermal Cells

AbstractLEAFY (LFY)-like proteins are plant-specific transcription factors that play essential roles in plant growth and development. In this study, a LEAFY homologue in soybean [Glycine max (L.) Merr.], designated as GmLFY, was cloned from inflorescences by the rapid amplification of cDNA ends (RACE) method. Sequence analysis showed that GmLFY cDNA contained a 1221 bp open reading frame, encoding 407 amino acid residues with typical characteristics of transcription factors. Subcellular localization in onion epidermal cells indicated that GmLFY protein was located in the nucleus. Analysis of the GmLFY genomic structure showed that the GmLFY gene has two introns with similar spliced sites as LFY-like genes in other plants. Reverse transcription-polymerase chain reaction (RT-PCR) analysis indicated that GmLFY was predominantly expressed in reproductive organs, such as inflorescences, pods and developing seeds. Further analysis showed that the expression level of GmLFY was higher in the middle stage than in the early or late stages of seed development, suggesting an essential regulatory role of GmLFY in soybean seed development.

Molecular Cloning and Characterization of a Novel Dehydrin Gene from Ginkgo biloba

2006 ◽  
Vol 26 (3) ◽  
pp. 203-215 ◽  
Author(s):  
Zhongxiang Deng ◽  
Yiding Wang ◽  
Keji Jiang ◽  
Xuefen Liu ◽  
Weisheng Wu ◽  
...  
Keyword(s):  
Ginkgo Biloba ◽  
Genomic Sequence ◽  
Gene Promoter ◽  
Single Copy ◽  
Pcr Analysis ◽  
Amino Acid Residues ◽  
Living Fossil ◽  
Reading Frame ◽  
Shared Identity ◽  
Rapid Amplification

A full-length cDNA encoding a dehydrin was cloned from the living fossil plant Ginkgo biloba by rapid amplification of cDNA ends (RACE). The cDNA, designated as GbDHN, was 813 bp long containing an open reading frame of 489 bp. The deduced GbDHN protein had 163 amino acid residues, which formed a 17 kDa polypeptide with a predicted isoelectric point (pI) of 5.75. GbDHN had an S-segment and a K-segment, indicative of dehydrins, but no Y-segments. Homology analysis indicated that the S-segment and K-segment of GbDHN shared identity with those of other reported dehydrins, indicating that GbDHN belonged to dehydrin superfamily. Genomic sequence of GbDHN was also cloned using genomic walker technology. By comparing genomic DNA with the cDNA, it was found that there was a 257-bp intron in this gene. Promoter analysis indicated that it contained six CAAT boxes, one TATA box, one ABRE box and one GC-motif in the 5′-flanking region. Southern blot analysis revealed that GbDHN belonged to a single copy gene family. RT-PCR analysis revealed that GbDHN constitutively expressed in stems and roots. The increased expression of GbDHN was detected when G. biloba seedlings were treated with exogenous abscisic acid (ABA), salt stress and drought stress. These results indicate that the GbDHN has the potential to play a role in response to ABA and environmental stresses that can cause plant dehydration.

scholarly journals Molecular cloning, expression and characterization of Bmserpin-2 gene from Bombyx mori.

2009 ◽  
Vol 56 (4) ◽  
Author(s):  
Ye Pan ◽  
Hengchuan Xia ◽  
Peng Lü ◽  
KePing Chen ◽  
Qin Yao ◽  
...  
Keyword(s):  
Bombyx Mori ◽  
Developmental Stages ◽  
Pcr Analysis ◽  
Post Inoculation ◽  
Amino Acid Residues ◽  
Gene Encoding ◽  
Reading Frame ◽  
E Coli ◽  
Real Time Quantitative Pcr ◽  
Localization Analysis

Serpins are a broadly distributed family of protease inhibitors. In this study, the gene encoding Bombyx mori serpin-2 (Bmserpin-2) was cloned and expressed in E. coli. The Bmserpin-2 cDNA contains a 1125 bp open reading frame (ORF). The deduced protein has 374 amino-acid residues, contains a conserved SERPIN domain and shares extensive homology with other invertebrate serpins. RT-PCR analysis showed that Bmserpin-2 was expressed in all developmental stages of B. mori larvae and various larval tissues. Subcellular localization analysis indicated that Bmserpin-2 protein was located in the cytoplasm. Interestingly, real-time quantitative PCR revealed that the expression of Bmserpin-2 in the midgut of susceptible B. mori strain 306 significantly increased at 72 hours post inoculation (hpi) when infected with BmNPV. However, there was no significant increase of the Bmserpin-2 expression in resistant strain NB infected with BmNPV. Thus, our data indicates that Bmserpin-2 may be involved in B. mori antiviral response.

Molecular cloning and assessment of the immunocontraceptive potential of the zona pellucida subunit 3 from Brandt's vole (Microtus brandti)

2006 ◽  
Vol 18 (3) ◽  
pp. 331 ◽  
Author(s):  
Hui Li ◽  
Yun-shang Piao ◽  
Zhi-bin Zhang ◽  
Christopher M. Hardy ◽  
Lyn A. Hinds
Keyword(s):  
Amino Acid ◽  
Zona Pellucida ◽  
Keyhole Limpet Hemocyanin ◽  
Amino Acid Residues ◽  
Igg Antibody ◽  
Reading Frame ◽  
Rapid Amplification ◽  
Ovarian Pathology ◽  
Race Pcr ◽  
Brandt’S Vole

A full-length cDNA encoding Brandt’s vole (Microtus brandti) zona pellucida glycoprotein subunit 3 (vZP3) was isolated using rapid amplification of cDNA ends–polymerase chain reaction (RACE-PCR). The cDNA contains an open reading frame of 1254 nucleotides encoding a polypeptide of 418 amino acid residues. The deduced amino acid sequence of vZP3 revealed high overall homology with hamster (82.1%), mouse (81.3%) and rat (80.6%). A synthetic vZP3 peptide corresponding to amino acid residues 328–343 was conjugated to keyhole limpet hemocyanin (KLH-vZP3328–343) and used to immunise female Brandt’s voles in order to test the efficacy of this peptide as a contraceptive antigen. High IgG antibody levels to the vZP3328–343 peptide were present in the sera of female voles immunised with KLH-vZP3328–343 and these also cross-reacted to the zona pellucida in ovaries of Brandt’s vole. The fertility of the KLH-vZP3328–343-immunised voles was reduced by 50% compared with controls without evidence of significant ovarian pathology.

Molecular cloning and characterization of cold-responsive gene Cbrci35 from Capsella bursa-pastoris

Biologia ◽  
2007 ◽  
Vol 62 (6) ◽  
Author(s):  
Juan Lin ◽  
Wen Zhang ◽  
Xuanwei Zhou ◽  
Xinglong Wang ◽  
Mingzhu Shi ◽  
...  
Keyword(s):  
Cold Acclimation ◽  
Leucine Zipper ◽  
Cell Attachment ◽  
Pcr Analysis ◽  
Sequence Motif ◽  
Amino Acid Residues ◽  
Cold Response ◽  
Rapid Amplification ◽  
High Level

AbstractA new rare cold-inducible (RCI) gene designated Cbrci35 was cloned from Capsella bursa-pastoris, an edible wild herb, using the rapid amplification of cDNA ends (RACE) method. The full-length cDNA of Cbrci35 (Database Accession No.: AY566573) was 1300 bp and contained a 978 bp ORF encoding a precursor of 326 amino acid residues with a 23 amino acids signal peptide. The predicted Cbrci35 protein contained a peroxidase active site and proximal heme-ligand signatures, an RGD cell attachment sequence motif and two leucine zipper pattern motifs. Bioinformatics analysis revealed that Cbrci35 has a high level of similarity with RCI genes from Arabidopsis thaliana and peroxidases genes from other plants. RT-PCR analysis revealed that Cbrci35 expressed only in root. A cold acclimation assay showed that Cbrci35 was expressed immediately after cold triggering, but this expression was transient, suggesting that it concerns cold acclimation. But expression was not induced exposed to dehydration, salt stress or abscisic acid, indicating that it might be subjected specifically to cold regulation. These results indicate that Cbrci35 is an analogue of RCI genes and may participate in cold-response or increasing the freezing tolerance of plants.

scholarly journals Combinatorial interactions of the LEC1 transcription factor specify diverse developmental programs during soybean seed development

2019 ◽  
Vol 117 (2) ◽  
pp. 1223-1232 ◽  
Author(s):  
Leonardo Jo ◽  
Julie M. Pelletier ◽  
Ssu-Wei Hsu ◽  
Russell Baden ◽  
Robert B. Goldberg ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Transcription Factors ◽  
Seed Development ◽  
Target Genes ◽  
Soybean Seed ◽  
Sequence Motifs ◽  
Dna Motifs ◽  
Regulatory Modules ◽  
Gene Sets ◽  
Combinatorial Interactions

The LEAFY COTYLEDON1 (LEC1) transcription factor is a central regulator of seed development, because it controls diverse biological programs during seed development, such as embryo morphogenesis, photosynthesis, and seed maturation. To understand how LEC1 regulates different gene sets during development, we explored the possibility that LEC1 acts in combination with other transcription factors. We identified and compared genes that are directly transcriptionally regulated by ABA-RESPONSIVE ELEMENT BINDING PROTEIN3 (AREB3), BASIC LEUCINE ZIPPER67 (bZIP67), and ABA INSENSITIVE3 (ABI3) with those regulated by LEC1. We showed that LEC1 operates with specific sets of transcription factors to regulate different gene sets and, therefore, distinct developmental processes. Thus, LEC1 controls diverse processes through its combinatorial interactions with other transcription factors. DNA binding sites for the transcription factors are closely clustered in genomic regions upstream of target genes, defining cis-regulatory modules that are enriched for DNA sequence motifs that resemble sequences known to be bound by these transcription factors. Moreover, cis-regulatory modules for genes regulated by distinct transcription factor combinations are enriched for different sets of DNA motifs. Expression assays with embryo cells indicate that the enriched DNA motifs are functional cis elements that regulate transcription. Together, the results suggest that combinatorial interactions between LEC1 and other transcription factors are mediated by cis-regulatory modules containing clustered cis elements and by physical interactions that are documented to occur between the transcription factors.

WRKY transcription factors actively respond to Fusarium oxysporum in Lilium regale Wilson

2021 ◽  
Author(s):  
Shan Li ◽  
Guanze Liu ◽  
Limei Pu ◽  
Xuyan Liu ◽  
Zie Wang ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Fusarium Oxysporum ◽  
Stress Responses ◽  
Expression Patterns ◽  
Pcr Analysis ◽  
Wrky Transcription Factors ◽  
Lilium Regale ◽  
Acid Methyl ◽  
Onion Epidermal Cells ◽  
Lilium Regale Wilson

The WRKY transcription factors (TFs) form a plant-specific superfamily important for regulating plant development, stress responses, and hormone signal transduction. In this study, many WRKY genes (LrWRKY1–35) were identified in Lilium regale Wilson, which is a wild lily species highly resistant to Fusarium wilt. These WRKY genes were divided into three classes (I–III) based on a phylogenetic analysis. The Class II WRKY TFs were further divided into five subclasses (IIa, IIb, IIc, IId, and IIe). Moreover, the gene expression patterns based on a quantitative real-time PCR analysis revealed the WRKY genes were differentially expressed in the L. regale roots, stems, leaves, and flowers. Additionally, the expression of the WRKY genes was affected by an infection by Fusarium oxysporum as well as by salicylic acid, methyl jasmonate, ethephon, and hydrogen peroxide treatments. Moreover, the LrWRKY1 protein was localized to the nucleus of onion epidermal cells. The recombinant LrWRKY1 protein purified from Escherichia coli bound specifically to DNA fragments containing the W-box sequence, and a yeast one-hybrid assay indicated that LrWRKY1 can activate transcription. A co-expression assay in tobacco confirmed LrWRKY1 regulates the expression of LrPR10-5. Furthermore, the overexpression of LrWRKY1 in tobacco and the Oriental Hybrid ‘Siberia’ (susceptible to F. oxysporum) increased the resistance of the transgenic plants to F. oxysporum. Overall, LrWRKY1 regulates the expression of the resistance gene LrPR10-5 and is involved in the defense response of L. regale to F. oxysporum. This study provides valuable information regarding the expression and functional characteristics of L. regale WRKY genes.

scholarly journals Molecular Cloning, Structure and Phylogenetic Analysis of a Hemocyanin Subunit from the Black Sea Crustacean Eriphia verrucosa (Crustacea, Malacostraca)

Genes ◽  
2021 ◽  
Vol 12 (1) ◽  
pp. 93
Author(s):  
Elena Todorovska ◽  
Martin Ivanov ◽  
Mariana Radkova ◽  
Alexandar Dolashki ◽  
Pavlina Dolashka
Keyword(s):  
Phylogenetic Analysis ◽  
Black Sea ◽  
The Black Sea ◽  
Copper Binding ◽  
Amino Acid Residues ◽  
Panulirus Interruptus ◽  
Reading Frame ◽  
Est Analysis ◽  
Physiological Processes ◽  
Rapid Amplification

Hemocyanins are copper-binding proteins that play a crucial role in the physiological processes in crustaceans. In this study, the cDNA encoding hemocyanin subunit 5 from the Black sea crab Eriphia verrucosa (EvHc5) was cloned using EST analysis, RT-PCR and rapid amplification of the cDNA ends (RACE) approach. The full-length cDNA of EvHc5 was 2254 bp, consisting of a 5′ and 3′ untranslated regions and an open reading frame of 2022 bp, encoding a protein consisting of 674 amino acid residues. The protein has an N-terminal signal peptide of 14 amino acids as is expected for proteins synthesized in hepatopancreas tubule cells and secreted into the hemolymph. The 3D model showed the presence of three functional domains and six conserved histidine residues that participate in the formation of the copper active site in Domain 2. The EvHc5 is O-glycosylated and the glycan is exposed on the surface of the subunit similar to Panulirus interruptus. The phylogenetic analysis has shown its close grouping with γ-type of hemocyanins of other crustacean species belonging to order Decapoda, infraorder Brachyura.

The alcohol dehydrogenase with a broad range of substrate specificity regulates vitality and reproduction of the plant-parasitic nematodeBursaphelenchus xylophilus

Parasitology ◽  
2018 ◽  
Vol 146 (4) ◽  
pp. 497-505 ◽  
Author(s):  
Linsong Wang ◽  
Tingting Zhang ◽  
Zhengsong Pan ◽  
Lulu Lin ◽  
Guoqing Dong ◽  
...  
Keyword(s):  
Alcohol Dehydrogenase ◽  
Inclusion Bodies ◽  
Bacterial Species ◽  
Pine Wilt Disease ◽  
Bursaphelenchus Xylophilus ◽  
Pcr Analysis ◽  
Amino Acid Residues ◽  
Reading Frame ◽  
Expression Of Genes ◽  
Relative Molecular Weight

AbstractPine wilt disease, which is caused by the pine wood nematode (PWN),Bursaphelenchus xylophilus, has caused huge damage to pine forests around the world. In this study, we analysed the PWN transcriptome to investigate the expression of genes related to the associated bacterial speciesPseudomonas fluorescensand found that the geneadh-1 encoding alcohol dehydrogenase (ADH) was upregulated. The open reading frame ofadh-1, which encoded a protein of 352 amino acid residues, was cloned fromB. xylophilus. Recombinant ADH with a relative molecular weight of 39 kDa, was present mainly in inclusion bodies and was overexpressed inEscherichia coliBL21 (DE3) and purified after refolding. The biochemical assay revealed that recombinant ADH could catalyse the dehydrogen reaction of eight tested alcohols including ethanol in the presence of NAD+. Quantitative real-time RT-PCR analysis indicated that ethanol upregulatedadh-1 expression in PWN. Results of RNA interference and inhibition of ADH treatment indicated that downregulating expression ofadh-1 or inhibition of ADH could reduce ethanol tolerance and the vitality and reproduction ability ofB. xylophilus, suggesting thatadh-1 is involved in pathogenicity of PWN.

scholarly journals Analysis of miRNAs Targeted Storage Regulatory Genes during Soybean Seed Development Based on Transcriptome Sequencing

Genes ◽  
2019 ◽  
Vol 10 (6) ◽  
pp. 408 ◽  
Author(s):  
Jing-Yao Yu ◽  
Zhan-Guo Zhang ◽  
Shi-Yu Huang ◽  
Xue Han ◽  
Xin-Yu Wang ◽  
...  
Keyword(s):  
Expression Pattern ◽  
Seed Development ◽  
Small Rna ◽  
Target Genes ◽  
Soybean Seed ◽  
Regulatory Genes ◽  
Small Rna Sequencing ◽  
Grain Development ◽  
Regulatory Relationship ◽  
Novel Mirna

Soybeans are an important cash crop and are widely used as a source of vegetable protein and edible oil. MicroRNAs (miRNA) are endogenous small RNA that play an important regulatory role in the evolutionarily conserved system of gene expression. In this study, we selected four lines with extreme phenotypes, as well as high or low protein and oil content, from the chromosome segment substitution line (CSSL) constructed from suinong (SN14) and ZYD00006, and planted and sampled at three stages of grain development for small RNA sequencing and expression analysis. The sequencing results revealed the expression pattern of miRNA in the materials, and predicted miRNA-targeted regulatory genes, including 1967 pairs of corresponding relationships between known-miRNA and their target genes, as well as 597 pairs of corresponding relationships between novel-miRNA and their target genes. After screening and annotating genes that were targeted for regulation, five specific genes were identified to be differentially expressed during seed development and subsequently analyzed for their regulatory relationship with miRNAs. The expression pattern of the targeted gene was verified by Real-time Quantitative PCR (RT-qPCR). Our research provides more information about the miRNA regulatory network in soybeans and further identifies useful genes that regulate storage during soy grain development, providing a theoretical basis for the regulation of soybean quality traits.

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