Cloning, characterization and expression analysis of a 1-aminocyclopropane-1-carboxylate synthase gene from pear

2013 ◽  
Vol 93 (3) ◽  
pp. 465-471
Author(s):  
Haiyan Shi ◽  
Yuxing Zhang ◽  
Liang Chen
Keyword(s):  
Gene Product ◽  
Expression Analysis ◽  
Cdna Clone ◽  
Phylogenetic Analyses ◽  
Pcr Amplification ◽  
Acc Synthase ◽  
Pcr Analysis ◽  
Pyrus Pyrifolia ◽  
Reading Frame ◽  
Amplification Technique

Shi, H., Zhang, Y. and Chen, L. 2013. Cloning, characterization and expression analysis of a 1-aminocyclopropane-1-carboxylate synthase gene from pear. Can. J. Plant Sci. 93: 465–471. In this study, a cDNA clone encoding putative 1-aminocyclopropane-1-carboxylate (ACC) synthase (ACS) that catalyzes the conversion of S-adenosyl-L-methionine to ACC in ethylene biosynthetic pathway was isolated from a cDNA library produced using mRNA from pear (Pyrus pyrifolia). The cDNA clone, designated PpACS2, comprised an open reading frame of 1, 341 bp encoding a protein of 446 amino acids that shares high similarity with the known plant ACSs. Using PCR amplification technique, a genomic clone (GenBank accession number: KC146402) corresponding to PpACS2 was isolated and shown to contain two introns. The PpACS2 gene product shared 97% identity with an ACC synthase from pear (Pyrus communis). Phylogenetic analyses clearly placed the gene product in the ACC synthase cluster of plant ACS superfamily tree. Quantitative RT-PCR analysis indicated that the PpACS2 gene was preferentially expressed in young pear leaves and shoots. The transcript of PpACS2 gene was accumulated at relatively high levels in anthers, but no signal was detected in the petals and mesocarp of pear. These results suggest that the PpACS2 may participate in the regulation of ethylene production in pear leaves, shoots, and anthers.

Molecular characterization of pear 14-3-3b gene regulated during fruit development

2016 ◽  
Vol 96 (3) ◽  
pp. 433-438
Author(s):  
Haiyan Shi ◽  
Yujing Zhao ◽  
Xuemin An ◽  
Yuxing Zhang
Keyword(s):  
Protein Interactions ◽  
Pcr Amplification ◽  
Pcr Analysis ◽  
Pyrus Pyrifolia ◽  
Biological Processes ◽  
Protein Protein Interactions ◽  
Rt Pcr ◽  
Developmentally Regulated ◽  
Amplification Technique

Plant 14-3-3 proteins (14-3-3s) are known to function in protein–protein interactions that mediate signal transduction pathways regulating many biological processes. The cDNA encoding putative 14-3-3 protein was isolated from pear (Pyrus pyrifolia) and designated Pp14-3-3b. Using the PCR amplification technique, the genomic clone corresponding to Pp14-3-3b was isolated and shown to contain six introns. Phylogenetic analysis clearly demonstrated that Pp14-3-3b was classified into the non-ɛ class of 14-3-3 superfamilies. Quantitative RT-PCR analysis indicated that the expression of the Pp14-3-3b gene was developmentally regulated in fruit. This study suggested that Pp14-3-3b might be involved in fruit ripening and the senescence of pear.

Pear IAA1 gene encoding an auxin-responsive Aux/IAA protein is involved in fruit development and response to salicylic acid

2014 ◽  
Vol 94 (2) ◽  
pp. 263-271 ◽  
Author(s):  
Haiyan Shi ◽  
Yanhui Wang ◽  
Zhenghong Li ◽  
Diansheng Zhang ◽  
Yufeng Zhang ◽  
...  
Keyword(s):  
Salicylic Acid ◽  
Fruit Development ◽  
Phylogenetic Analyses ◽  
Pcr Amplification ◽  
Pyrus Pyrifolia ◽  
Dimerization Domain ◽  
Developmentally Regulated ◽  
Gene Encoding ◽  
Domain Profile ◽  
Localization Signal

Shi, H., Wang, Y., Li, Z., Zhang, D., Zhang, Y., Xiang, D., Li, Y. and Zhang, Y. 2014. Pear IAA1 gene encoding an auxin-responsive Aux/IAA protein is involved in fruit development and response to salicylic acid. Can. J. Plant Sci. 94: 263–271. Auxin-responsive Aux/IAA proteins are rapidly auxin-induced, short-lived proteins that act as repressors for the auxin response factor (ARF)-activated gene expression. A gene encoding an Aux/IAA protein and designated PpIAA1 was isolated from pear (Pyrus pyrifolia). Using PCR amplification techniques, the genomic clone corresponding to PpIAA1 was isolated and shown to contain three introns with typical GT/AG boundaries defining the splice junctions. The deduced PpIAA1 protein contains the conserved features of indole-3-acetic acids (IAA): four Aux/IAA conserved domains, Aux/IAA family domain, Aux/IAA-ARF dimerization domain profile, and conserved nuclear localization signal (NLS) motifs. Phylogenetic analyses clearly demonstrated PpIAA1 has the highest homology with grape VvIAA. PpIAA1 was preferentially expressed in fruit, and moderate expression was found in anthers. Relatively low expression signal was detected in other tissues including shoots, leaves, and petals. Moreover, expression of PpIAA1 was developmentally regulated in fruit. Further study demonstrated that PpIAA1 expression in pear fruit was remarkably regulated by salicylic acid and IAA. The data suggest that PpIAA1 might be involved in the interplay between IAA and salicylic acid signaling pathway during the fruit development of pear.

scholarly journals Molecular cloning of RhD cDNA derived from a gene present in RhD- positive, but not RhD-negative individuals

Blood ◽  
1993 ◽  
Vol 82 (2) ◽  
pp. 651-655 ◽  
Author(s):  
MA Arce ◽  
ES Thompson ◽  
S Wagner ◽  
KE Coyne ◽  
BA Ferdman ◽  
...  
Keyword(s):  
Cdna Clone ◽  
Molecular Genetic ◽  
Pcr Amplification ◽  
Pcr Analysis ◽  
Blood Group System ◽  
Pcr Cloning ◽  
Antigenic Protein ◽  
Genomic Libraries ◽  
Polymerase Chain ◽  
Rh Blood Group

The Rh blood group system plays a major role in immune and nonimmune hemolytic states. Although an Rh cDNA has been previously cloned, there is no information on which Rh antigenic protein it encodes. Using polymerase chain reaction (PCR) amplification, we have identified this original Rh clone, here designated Rh21, and an additional Rh cDNA clone, Rh13, that is 96% nucleotide- and 92% amino acid-identical to Rh21, with the substitutions scattered throughout the sequence. A molecular genetic approach was used to match this Rh clone with an Rh specificity. The mRNA transcript for Rh13 was present in reticulocytes from RhD-positive individuals, but was absent from the reticulocytes of RhD-negative individuals. Using conventional screening of genomic libraries, as well as PCR cloning, partial genomic clones for these two Rh cDNAs were obtained. Based on PCR analysis and Southern blots, the Rh21 gene was present in all individuals, but an intact Rh13 gene was only present in RhD-positive and not RhD-negative individuals. Thus, by correlating the presence of Rh mRNA and gene sequences with individual Rh phenotypes, we were able to establish that the new Rh13 cDNA clone represents the RhD protein.

scholarly journals Molecular cloning of RhD cDNA derived from a gene present in RhD- positive, but not RhD-negative individuals

Blood ◽  
1993 ◽  
Vol 82 (2) ◽  
pp. 651-655 ◽  
Author(s):  
MA Arce ◽  
ES Thompson ◽  
S Wagner ◽  
KE Coyne ◽  
BA Ferdman ◽  
...  
Keyword(s):  
Cdna Clone ◽  
Molecular Genetic ◽  
Pcr Amplification ◽  
Pcr Analysis ◽  
Blood Group System ◽  
Pcr Cloning ◽  
Antigenic Protein ◽  
Genomic Libraries ◽  
Polymerase Chain ◽  
Rh Blood Group

Abstract The Rh blood group system plays a major role in immune and nonimmune hemolytic states. Although an Rh cDNA has been previously cloned, there is no information on which Rh antigenic protein it encodes. Using polymerase chain reaction (PCR) amplification, we have identified this original Rh clone, here designated Rh21, and an additional Rh cDNA clone, Rh13, that is 96% nucleotide- and 92% amino acid-identical to Rh21, with the substitutions scattered throughout the sequence. A molecular genetic approach was used to match this Rh clone with an Rh specificity. The mRNA transcript for Rh13 was present in reticulocytes from RhD-positive individuals, but was absent from the reticulocytes of RhD-negative individuals. Using conventional screening of genomic libraries, as well as PCR cloning, partial genomic clones for these two Rh cDNAs were obtained. Based on PCR analysis and Southern blots, the Rh21 gene was present in all individuals, but an intact Rh13 gene was only present in RhD-positive and not RhD-negative individuals. Thus, by correlating the presence of Rh mRNA and gene sequences with individual Rh phenotypes, we were able to establish that the new Rh13 cDNA clone represents the RhD protein.

Pear PIP1 gene is regulated during fruit development and is invovled in response to salicylic acid and ethylene

2015 ◽  
Vol 95 (1) ◽  
pp. 77-85
Author(s):  
Haiyan Shi ◽  
Yanhui Wang ◽  
Diansheng Zhang ◽  
Liang Chen ◽  
Yuxing Zhang
Keyword(s):  
Salicylic Acid ◽  
Plasma Membrane ◽  
Fruit Development ◽  
Phylogenetic Analyses ◽  
Pcr Amplification ◽  
Protein Domain ◽  
Pyrus Pyrifolia ◽  
Plasma Membrane Intrinsic Protein ◽  
Intrinsic Protein ◽  
Young Leaves

Shi, H., Wang, Y., Zhang, D., Chen, L. and Zhang, Y. 2015. Pear PIP1 gene is regulated during fruit development and is invovled in response to salicylic acid and ethylene. Can. J. Plant Sci. 95: 77–85. Plasma membrane intrinsic proteins (PIPs), a subfamily of aquaporins, are widely implicated in plant growth and development. A gene encoding a plasma membrane intrinsic protein and designated PpPIP1 was isolated from pear (Pyrus pyrifolia). Using PCR amplification techniques, the genomic clone corresponding to PpPIP1 was isolated and shown to contain three introns with typical GT/AG boundaries defining the splice junctions. The deduced PpPIP1 protein contains the conserved features of PIPs: six transmembrane α-helices, a major intrinsic protein domain, and a conserved asparagine–proline–alanine (NPA) signature sequence. Phylogenetic analyses clearly demonstrated that PpPIP1 has the highest homology with apple (Malus×domestica) MdPIP1a and Malus hupehensis MhPIP1-1. PpPIP1 transcripts were mainly detected in young leaves, shoots, petals and mesocarp of fruit, but a relatively low expression signal was detected in anthers. In particular, expression of PpPIP1 was developmentally regulated in fruit. Further study demonstrated that PpPIP1 expression in pear fruit was down-regulated by salicylic acid (SA) and up-regulated by ethylene. These data suggest that PpPIP1 may be involved in the response to SA and ethylene during fruit development, which would provide valuable information for water permeability studies in pear.

scholarly journals Genome-Wide Identification and Transcriptional Expression Analysis of Cucumber Superoxide Dismutase (SOD) Family in Response to Various Abiotic Stresses

2017 ◽  
Vol 2017 ◽  
pp. 1-14 ◽  
Author(s):  
Yong Zhou ◽  
Lifang Hu ◽  
Hao Wu ◽  
Lunwei Jiang ◽  
Shiqiang Liu
Keyword(s):  
Superoxide Dismutase ◽  
Expression Analysis ◽  
Stress Responses ◽  
Phylogenetic Analyses ◽  
Promoter Sequence ◽  
Pcr Analysis ◽  
Motif Analysis ◽  
Genome Wide ◽  
Potential Applications ◽  
Almost All

Superoxide dismutase (SOD) proteins are widely present in the plant kingdom and play important roles in different biological processes. However, little is known about the SOD genes in cucumber. In this study, night SOD genes were identified from cucumber (Cucumis sativus) using bioinformatics-based methods, including 5 Cu/ZnSODs, 3 FeSODs, and 1 MnSOD. Gene structure and motif analysis indicated that most of the SOD genes have relatively conserved exon/intron arrangement and motif composition. Phylogenetic analyses with SODs from cucumber and several other species revealed that these SOD proteins can be traced back to two ancestral SODs before the divergence of monocot and dicot plants. Many cis-elements related to stress responses and plant hormones were found in the promoter sequence of each CsSOD gene. Gene expression analysis revealed that most of the CsSOD genes are expressed in almost all the tested tissues. qRT-PCR analysis of 8 selected CsSOD genes showed that these genes could respond to heat, cold, osmotic, and salt stresses. Our results provide a basis for further functional research on SOD gene family in cucumber and facilitate their potential applications in the genetic improvement of cucumber.

scholarly journals Genome-Wide Identification and Expression Analysis of Flavonoid 3'-Hydroxylase Gene Family in Carthamus Tinctorius L.

2021 ◽  
Author(s):  
Hoàng Việt Nguyễn Quốc ◽  
Kong Jie ◽  
Naveed Ahmad ◽  
Yang Zhuoda ◽  
Wang Nan ◽  
...  
Keyword(s):  
Expression Analysis ◽  
Phylogenetic Analyses ◽  
Evolutionary Relationship ◽  
Pcr Analysis ◽  
Genetic Evolution ◽  
Significant Information ◽  
Protein Motifs ◽  
Qrt Pcr ◽  
Digital Expression ◽  
Genome Wide

Abstract ObjectiveThrough experiments and bioinformatic analysis clearly demonstrate considerable information about the genetic evolution of the flavonoid 3'-hydroxylase (F3'H) gene in Safflower and in plants.ResultsHere, we performed genome wide survey of safflower genome and identified a total of 22 CtF3'H enzyme encoding genes. Phylogenetic analyses revealed the classifications of these CtF3'Hs into nine subgroups demonstrating their evolutionary relationship. The distribution of the conserved protein motifs, and cis-regulatory units of CtF3'Hs indicated essential structure-to-function components leading to the final function of protein or its interactions. Furthermore, the results of digital expression analysis and the qRT-PCR pattern of 22 putative CtF3’H genes during different flowering stages suggested their requisite roles in safflower petal pigmentation. In addition, the fusions construct of plant expression vector pCAMBIA1302-GFP-CtF3’H5 in onion epidermal cells verified the subcellular localization of CtF3’H5 to the plasma membrane. Subsequently, the prokaryotic expression and western blot hybridization of CtF3’H5 resulted in a stable 50.3kD target protein. These results partly demonstrate the influence of F3'Hs on plants.ConclusionsIn this study, the results of digital expression and qRT-PCR analysis of 22 putative CtF3'H genes in different flowering stages indicate their essential role in safflower petal pigmentation. clearly demonstrates significant information on the genetic evolution of important enzyme-coding genes and will provide a pathway for future functional studies of F3'Hs in safflower as well as in plants.

scholarly journals Molecular Epidemiology and Whole-Genome Analysis of Bovine Foamy Virus in Japan

Viruses ◽  
2021 ◽  
Vol 13 (6) ◽  
pp. 1017
Author(s):  
Hirohisa Mekata ◽  
Tomohiro Okagawa ◽  
Satoru Konnai ◽  
Takayuki Miyazawa
Keyword(s):  
Phylogenetic Analyses ◽  
Foamy Virus ◽  
Pcr Analysis ◽  
Whole Genome ◽  
Genome Sequences ◽  
Whole Genome Analysis ◽  
Virus Family ◽  
Bovine Foamy Virus ◽  
Novel Genotype

Bovine foamy virus (BFV) is a member of the foamy virus family in cattle. Information on the epidemiology, transmission routes, and whole-genome sequences of BFV is still limited. To understand the characteristics of BFV, this study included a molecular survey in Japan and the determination of the whole-genome sequences of 30 BFV isolates. A total of 30 (3.4%, 30/884) cattle were infected with BFV according to PCR analysis. Cattle less than 48 months old were scarcely infected with this virus, and older animals had a significantly higher rate of infection. To reveal the possibility of vertical transmission, we additionally surveyed 77 pairs of dams and 3-month-old calves in a farm already confirmed to have BFV. We confirmed that one of the calves born from a dam with BFV was infected. Phylogenetic analyses revealed that a novel genotype was spread in Japan. In conclusion, the prevalence of BFV in Japan is relatively low and three genotypes, including a novel genotype, are spread in Japan.

scholarly journals Cloning, Characterization, and RNA Interference Effect of the UDP-N-Acetylglucosamine Pyrophosphorylase Gene in Cnaphalocrocis medinalis

Genes ◽  
2021 ◽  
Vol 12 (4) ◽  
pp. 464
Author(s):  
Yuan-Jin Zhou ◽  
Juan Du ◽  
Shang-Wei Li ◽  
Muhammad Shakeel ◽  
Jia-Jing Li ◽  
...  
Keyword(s):  
Developmental Disorders ◽  
Developmental Stages ◽  
Phylogenetic Analyses ◽  
Digital Pcr ◽  
Cnaphalocrocis Medinalis ◽  
Chitin Synthesis ◽  
Reading Frame ◽  
Rnai Technology ◽  
Glyphodes Pyloalis ◽  
Key Enzyme

The rice leaf folder, Cnaphalocrocis medinalis is a major pest of rice and is difficult to control. UDP-N-acetylglucosamine pyrophosphorylase (UAP) is a key enzyme in the chitin synthesis pathway in insects. In this study, the UAP gene from C. medinalis (CmUAP) was cloned and characterized. The cDNA of CmUAP is 1788 bp in length, containing an open reading frame of 1464 nucleotides that encodes 487 amino acids. Homology and phylogenetic analyses of the predicted protein indicated that CmUAP shared 91.79%, 87.89%, and 82.75% identities with UAPs of Glyphodes pyloalis, Ostrinia furnacalis, and Heortia vitessoides, respectively. Expression pattern analyses by droplet digital PCR demonstrated that CmUAP was expressed at all developmental stages and in 12 tissues of C. medinalis adults. Silencing of CmUAP by injection of double-stranded RNA specific to CmUAP caused death, slow growth, reduced feeding and excretion, and weight loss in C. medinalis larvae; meanwhile, severe developmental disorders were observed. The findings suggest that CmUAP is essential for the growth and development of C. medinalis, and that targeting the CmUAP gene through RNAi technology can be used for biological control of this insect.

scholarly journals Biological and Molecular Characteristics of a Novel Partitivirus Infecting the Edible Fungus Lentinula edodes

Plant Disease ◽  
2017 ◽  
Vol 101 (5) ◽  
pp. 726-733 ◽  
Author(s):  
Mengpei Guo ◽  
Yinbing Bian ◽  
Jinjie Wang ◽  
Gangzheng Wang ◽  
Xiaolong Ma ◽  
...  
Keyword(s):  
Lentinula Edodes ◽  
Control Strategies ◽  
Phylogenetic Analyses ◽  
Pcr Analysis ◽  
Molecular Characteristics ◽  
Rt Pcr ◽  
Edible Fungus ◽  
Qpcr Analysis ◽  
Disease Diagnostics ◽  
Wild Strains

A new partitivirus named Lentinula edodes partitivirus 1 (LePV1) was isolated from a diseased L. edodes strain with severe degeneration of the mycelium and imperfect browning in bag cultures. The nucleotide sequences of LePV1 dsRNA-1 and dsRNA-2 were determined; they were 2,382 bp and 2,245 bp in length, and each contained a single ORF encoding RNA-dependent RNA polymerase (RdRp) and coat protein (CP), respectively. The purified virus preparation contained isometric particles 34 nm in diameter encapsidating these dsRNAs. Phylogenetic analyses showed LePV1 to be a new member of Betapartitivirus, with the RdRp sequence most closely related to Grapevine partitivirus. RT-PCR analysis showed that 27 of the 56 Chinese L. edodes core collection strains carry LePV1, with the virus being more common in wild strains than cultivated strains. In addition, qPCR analysis suggested that coinfection with L. edodes mycovirus HKB (LeV-HKB) could increase replication of the RdRp gene of LePV1. This study may be essential for the development of more accurate disease diagnostics and the formulation of control strategies for viral diseases in L. edodes.

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