Rapid Freeze Acclimation of Poncirus trifoliata Seedlings Exposed to 10 °C and Long Days

Poncirus trifoliata (L.) Raf. seeds were germinated in perlite under intermittent mist at about 25 °C and natural daylight in a greenhouse. Two-week-old seedlings were then transferred into a growth chamber at 25 °C and 16-hour daylength for 1 week. Tissue samples were collected at 0, 6, 24, 168, and 504 hours after temperature equilibration at 10 °C. Freezing tolerance at –6.7 °C, as determined by electrolyte leakage, and stem (leaves attached) water potential (ψx), measured using a pressure chamber, was recorded for a subset of seedlings for each time interval. Red coloration (apparently anthocyanin) developed at the petiole leaflet junction and buds after 48 hours at 10 °C and gradually occurred throughout the leaves during further exposure. Complementary DNA clones for phenylalanine ammonia lyase (PAL), 4-coumarate: coA ligase (4CL), and chalcone synthase (CHS) were used to probe RNA isolated from the leaves. No increase in steady-state messenger RNA level was detected. Increases in freeze hardiness occurred within 6 hours in the leaves, and continued for up to 1 week. Water potential initially decreased from –0.6 to –2.0 MPa after 6 hours, then returned to –0.6 MPa after 1 week. Thus, Poncirus trifoliata seedlings freeze-acclimate significantly after only 6 hours at 10 °C.

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Changes in Freezing Tolerance, Water Potential, and Gene Expression of Poncirus trifoliata `Rubidoux' Seedlings Exposed to Acclimating Low Temperatures and Long Days

HortScience ◽

10.21273/hortsci.31.4.580a ◽

1996 ◽

Vol 31 (4) ◽

pp. 580a-580

Author(s):

Milton E. Tignor ◽

John M. Davis ◽

Frederick S. Davies ◽

Wayne B. Sherman

Keyword(s):

Gene Expression ◽

Low Temperature ◽

Water Potential ◽

Freezing Tolerance ◽

Freeze Tolerance ◽

Poncirus Trifoliata ◽

Economic Losses ◽

Tissue Samples ◽

Coumarate:Coa Ligase ◽

Hardy Cross

Poncirus trifoliata is a comparatively hardy, cross compatible, and graft compliant relative of Citrus. The citrus industry in Florida has suffered immense economic losses due to freezes. Although much research has been done in citrus freeze hardiness, little work has been on the early induction of freeze tolerance by low temperature. Poncirus trifoliata `Rubidoux' seedlings were germinated in perlite under intermittent mist at about 25°C and natural daylight conditions in a greenhouse and grown 2 weeks. See dlings were then transferred into a growth chamber at 25°C and 16 hour daylength for 1 week. Temperature was lowered to 10°C and tissue samples were collected at 0, 6, 24, and 168 hours. Freezing tolerance, at –6.7°C as determined by electrolyte leakage, and stem (leaves attached) water potential, measured using a pressure bomb, were also recorded for a subset of seedlings for the above intervals. After exposure to low temperature for 48 hours a red coloration became visible at the petiole leaflet junction an d at the buds, with subsequent exposure to low temperature the coloration spread to the leaves. Clones for phenylalanine ammonia lyase (PAL), 4-coumarate:CoA ligase (4CL), and chlorophyll ab binding protein (CAB), and chalcone synthase (CHS) were used to probe RNA isolated from P. trifoliata. PAL and 4CL transcripts increased in response to the low temperature. Significant increases in freeze hardiness occurred within 6 hours in the leaves, and increases continued for up to one week. Water potential increased from –0.6 to –2.0 MPa after 6 hours, then returned to –0.6 MPa after 1 week. These data indicate that increases in freezing tolerance and changes in water potential and gene expression can be detected shortly after low temperature treatments are imposed on P. trifoliata seedlings.

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Transcriptome Sequencing and Chemical Analysis Reveal the Formation Mechanism of White Florets in Carthamus tinctorius L.

Plants ◽

10.3390/plants9070847 ◽

2020 ◽

Vol 9 (7) ◽

pp. 847

Author(s):

Tingyan Qiang ◽

Jiushi Liu ◽

Yuqing Dong ◽

Yinbo Ma ◽

Bengang Zhang ◽

...

Keyword(s):

Transcriptome Sequencing ◽

Phenylalanine Ammonia Lyase ◽

Chalcone Synthase ◽

Chemical Constituents ◽

Carthamus Tinctorius ◽

Cdna Libraries ◽

Chain Reaction ◽

Coa Ligase ◽

Pigment Biosynthesis ◽

Polymerase Chain

Carthamus tinctorius L. (safflower), an economic crop and herb, has been extensively studied for its diverse chemical constituents and pharmacological effects, but the mechanism of safflower pigments (SP) leading to different colors of florets has not been clarified. In the present study, we compared the contents of SP in two varieties of safflower with white and red florets, named Xinhonghua No. 7 (WXHH) and Yunhong No. 2 (RYH). The results showed the contents of SP in RYH were higher than WXHH. To investigate genes related to SP, we obtained six cDNA libraries of florets from the two varieties by transcriptome sequencing. A total of 225,008 unigenes were assembled and 40 unigenes related to safflower pigment biosynthesis were annotated, including 7 unigenes of phenylalanine ammonia-lyase (PAL), 20 unigenes of 4-coumarate-CoA ligase (4CL), 1 unigene of trans-cinnamate 4-monooxygenase (C4H), 7 unigenes of chalcone synthase (CHS), 4 unigenes of chalcone isomerase (CHI), and 1 unigene of flavanone 3-hydroxylase (F3H). Based on expression levels we selected 16 differentially expressed unigenes (DEGs) and tested them using reverse transcription-quantitative real-time polymerase chain reaction (RT-qPCR), which was consistent with the sequencing results. Consequently, we speculated that in WXHH, 3 PALs, 3 4CLs, 1 C4H, 1 CHS, and 1 CHI, which were down-regulated, and 1 F3H, which was up-regulated, may play a key role in the formation of white florets.

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Human beta-globin messenger RNA. III. Nucleotide sequences derived from complementary DNA.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(17)40156-6 ◽

1977 ◽

Vol 252 (14) ◽

pp. 5040-5053 ◽

Cited By ~ 7

Author(s):

C A Marotta ◽

J T Wilson ◽

B G Forget ◽

S M Weissman

Keyword(s):

Messenger Rna ◽

Nucleotide Sequences ◽

Beta Globin ◽

Complementary Dna

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Expression of Cannabinoid Receptors in Myometrium and its Correlation With Dysmenorrhea in Adenomyosis

Reproductive Sciences ◽

10.1177/1933719119833483 ◽

2019 ◽

Vol 26 (12) ◽

pp. 1618-1625 ◽

Cited By ~ 3

Author(s):

Xue Shen ◽

Hua Duan ◽

Sha Wang ◽

Wei Hong ◽

Yu-Yan Wang ◽

...

Keyword(s):

Messenger Rna ◽

Cannabinoid Receptors ◽

Pain Perception ◽

Cannabinoid Receptor ◽

Quantitative Polymerase Chain Reaction ◽

Premenopausal Women ◽

Type I ◽

Junctional Zone ◽

Tissue Samples ◽

Expression Levels

The myometrium, especially the junctional zone (JZ), is now well documented to have a role in the pathogenesis of adenomyosis. Cannabinoid receptors have been shown to participate in the establishment of endometriosis and its pain perception. However, its relation to adenomyosis has not been identified yet. The aim of this study was to investigate the expression of cannabinoid receptor type I (CB1) and type II (CB2) in myometrium of uteri with and without adenomyosis and determine the correlation between their levels and clinical parameters of adenomyosis. We collected tissue samples of JZ and the outer myometrium from 45 premenopausal women with adenomyosis and 34 women without adenomyosis. CB1 and CB2 messenger RNA (mRNA) and protein expression levels were evaluated by the use of Western blotting and real-time quantitative polymerase chain reaction from all samples. Clinical information on the severity of dysmenorrhea and other data were collected. We found both CB1 and CB2 mRNA and protein levels in women with adenomyosis were significantly higher than those of controls, and CB1 expression levels in JZ were positively correlated with the severity of dysmenorrhea. These data suggest that cannabinoid receptor CB1 may be involved in the pathogenesis of dysmenorrhea in adenomyosis and may be a potential therapeutic target.

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Complement Factor C3 Methylation and mRNA Expression Is Associated to BMI and Insulin Resistance in Obesity

Genes ◽

10.3390/genes9080410 ◽

2018 ◽

Vol 9 (8) ◽

pp. 410 ◽

Cited By ~ 7

Author(s):

Daniel Castellano-Castillo ◽

Isabel Moreno-Indias ◽

Jose Carlos Fernandez-Garcia ◽

Mercedes Clemente-Postigo ◽

Manuel Castro-Cabezas ◽

...

Keyword(s):

Insulin Resistance ◽

Dna Methylation ◽

Adipose Tissue ◽

Messenger Rna ◽

Density Lipoprotein ◽

Complement Factor ◽

Mrna Levels ◽

Model Assessment ◽

Tissue Samples ◽

Obese Class

Epigenetic marks, and especially DNA methylation, are becoming an important factor in obesity, which could help to explain its etiology and associated comorbidities. Adipose tissue, now considered as an important endocrine organ, produces complement system factors. Complement component 3 (C3) turns out to be an important protein in metabolic disorders, via either inflammation or the C3 subproduct acylation stimulating protein (ASP) which directly stimulates lipid storage. In this study, we analyze C3 DNA methylation in adipose tissue from subjects with a different grade of obesity. Adipose tissue samples were collected from subjects with a different degree of obesity determined by their body mass index (BMI) as: Overweight subjects (BMI ≥ 25 and <30), obese class 1/2 subjects (BMI ≥ 30 and <40) and obese class 3 subjects (BMI ≥ 40). C3 DNA methylation was measured for 7 CpGs by pyrosequencition using the Pyromark technology (Qiagen, Madrid Spain). C3 messenger RNA (mRNA) levels were analyzed by pre-designed Taqman assays (Applied biosystems, Foster City, CA, USA) and ASP/C3a was measured using a ELISA kit. The data were analyzed using the statistic package SPSS. C3 DNA methylation levels were lower in the morbid obese group. Accordingly, C3 methylation correlated negatively with BMI and leptin. However, C3 mRNA levels were more associated with insulin resistance, and positive correlations with insulin, glucose and homeostasis model assessment-estimated insulin resistance (HOMA-IR) existed. ASP correlated negatively with high density lipoprotein (HDL) cholesterol. C3 methylation levels were associated to adiposity variables, such as BMI and leptin, while the C3 mRNA levels were associated to glucose metabolism.

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Chromosomal localization of parsley 4-coumarate: CoA ligase genes by in situ hybridization with a complementary DNA

Plant Cell Reports ◽

10.1007/bf00716838 ◽

1989 ◽

Vol 8 (2) ◽

pp. 59-62

Author(s):

Pung-Ling Huang ◽

Klaus Hahlbrock ◽

Imre E. Somssich

Keyword(s):

In Situ Hybridization ◽

Chromosomal Localization ◽

Complementary Dna ◽

Coa Ligase

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Cloning and Sequencing cDNA Encoding for Rhoptry-2 Toxoplasma Gondii Tachyzoite Local Isolate

Indonesian Journal of Biotechnology ◽

10.22146/ijbiotech.7555 ◽

2015 ◽

Vol 10 (2) ◽

Author(s):

Wayan T. Artama ◽

Yulia Sari ◽

Didik Tulus Subekti ◽

Soenarwan Hery Poerwanto ◽

Jarot Subandono

Keyword(s):

Toxoplasma Gondii ◽

Recombinant Plasmid ◽

Messenger Rna ◽

Secretory Protein ◽

Gene Encoding ◽

Isolation System ◽

Complementary Dna ◽

E Coli ◽

Local Isolate ◽

Cell Target

Rhoptry protein belongs to an excretory and secretory antigens (ESAs) that play an important role during activepenetration of parasite into the cell target. This protein an able Toxoplasma gondii to actively penetrate targetedcell, meanwhile ESAs protein stimulates intracellular vacuole modification. It is, therefore, after the parasitesuccessfully enter the cell target then Granule (GRA) proteins are responsible for the formation of parasitophorusvacuole, which is protect the fusion with other intracellular compartments such as lysosomal vacuole. Consequently,this parasite is being able to survive and multiply at the cell target. The current study was aimed to clone andsequens cDNA encoding for ROP-2 of local isolated T. gondii tachizoite through DNA recombinant technique.Total ribonucleic acid (RNA) was isolated from tachyzoites of local isolated T. gondii that were grown up in Balb/c mice. Messenger RNA was isolated from total RNA using PolyAtract mRNA Isolation System. Messenger RNA wasused as a template for synthesis cDNA using Riboclone cDNA Synthesis System AMV-RT. EcoRI adaptor fromRiboclone EcoRI Adaptor Ligation System was added to Complementary DNA and than ligated to pUC19. Recombinantplasmid was transformed into E. coli (XL1-Blue). The transformed E. coli XL-1 Blue were plated on LB agarcontaining X-Gal, IPTG and ampicillin. Recombinant clones (white colony) were picked up and grown up in theLB medium at 37oC overnight. Expression of recombinant protein was analysed by immunoblotting in order toidentify cDNA recombinant wich is express ESA of T. gondii local isolate. Recombinant plasmid were isolatedusing alkalilysis method and were elektroforated in 1% agarose gel. The isolated DNA recombinant plasmid wascut using Eco RI and then sequenced through Big Dye Terminator Mix AB1 377A Sequencer using M13 Forward andM13 Reverse primers. The conclusion of this results showed that the recombinant clone was coding for excretoryand secretory protein which has molecular weight of 54 kDa. The DNA alignments of sequence from the clonedgene showed 97% homology with gene encoding for ROP-2 of T. gondii RH isolate.Keywords: Toxoplasma gondii, tachizoite, ESA, complementary DNA, ROP2

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Differential accumulation of plant defense gene transcripts in a compatible and an incompatible plant-pathogen interaction

Molecular and Cellular Biology ◽

10.1128/mcb.6.5.1615-1623.1986 ◽

1986 ◽

Vol 6 (5) ◽

pp. 1615-1623

Author(s):

J N Bell ◽

T B Ryder ◽

V P Wingate ◽

J A Bailey ◽

C J Lamb

Keyword(s):

Plant Defense ◽

Phenylalanine Ammonia Lyase ◽

Chalcone Synthase ◽

Defense Responses ◽

Infected Tissue ◽

Colletotrichum Lindemuthianum ◽

Specific Interactions ◽

Defense Gene ◽

Sequence Of Events ◽

Gene Transcripts

Phenylalanine ammonia-lyase and chalcone synthase catalyze the first reaction of phenylpropanoid biosynthesis and the first reaction of a branch pathway specific for flavonoid-isoflavonoid biosynthesis, respectively. These enzymes are key control elements in the synthesis of kievitone, phaseollin, and related isoflavonoid-derived phytoalexins. RNA blot hybridization with 32P-labeled cDNA sequences was used to demonstrate marked accumulation of phenylalanine ammonia-lyase and chalcone synthase mRNAs in excision-wounded hypocotyls of Phaseolus vulgaris L. (dwarf French bean) and during race-cultivar-specific interactions between hypocotyls of P. vulgaris and the partially biotrophic fungus Colletotrichum lindemuthianum, the causal agent of anthracnose. In an incompatible interaction (host resistant), early concomitant accumulation of phenylalanine ammonia-lyase and chalcone synthase mRNAs, localized mainly but not entirely in tissue adjacent to the site of infection, was observed prior to the onset of phytoalexin accumulation and expression of localized, hypersensitive resistance. In contrast, in a compatible interaction (host susceptible) there was no early accumulation of these transcripts; instead, there was a delayed widespread response associated with phytoalexin accumulation during attempted lesion limitation. Two-dimensional gel electrophoresis of [35S]methionine-labeled polypeptides synthesized in vitro by translation of isolated polysomal RNA demonstrated stimulation of the synthesis of characteristic sets of phenylalanine ammonia-lyase and chalcone synthase isopolypeptides in directly infected tissue and distant, hitherto uninfected tissue in both compatible and incompatible interactions. Our data show that specific accumulation of plant defense gene transcripts is a key early component in the sequence of events leading to expression of defense responses in wounded tissue and in infected tissue during race-cultivar-specific interactions and that an elicitation signal is transmitted intercellularly in response to infection.

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Expression of RNA-binding motif 10 is associated with advanced tumor stage and malignant behaviors of lung adenocarcinoma cancer cells

Tumor Biology ◽

10.1177/1010428317691740 ◽

2017 ◽

Vol 39 (3) ◽

pp. 101042831769174 ◽

Cited By ~ 4

Author(s):

Guofang Guan ◽

Ranwei Li ◽

Wenfang Tang ◽

Tiecheng Liu ◽

Zhenzhong Su ◽

...

Keyword(s):

Lung Adenocarcinoma ◽

Messenger Rna ◽

Rna Binding ◽

Tumor Stage ◽

Receptor Interaction ◽

Binding Motif ◽

Advanced Tumor Stage ◽

Complementary Dna ◽

Advanced Tumor ◽

Lung Adenocarcinoma Cancer

This study assessed RNA-binding motif 10 expression in lung adenocarcinoma tissues and examined the role and mechanism of RNA-binding motif 10 in the regulation of lung adenocarcinoma malignancy. Lung adenocarcinoma and corresponding adjacent non-tumor lung tissues from 41 patients were subjected to reverse transcription-polymerase chain reaction and Western blot assessment to detect RNA-binding motif 10 expression. Recombinant lentivirus carrying RNA-binding motif 10 complementary DNA was used to infect lung adenocarcinoma cell lines, A549 and H1299 cells. Complementary DNA microarray was used to profile RNA-binding motif 10–regulated genes. Levels of RNA-binding motif 10 messenger RNA and protein were significantly lower in lung adenocarcinoma tissues than those in paired non-tumor tissues (p < 0.001). Reduced RNA-binding motif 10 expression was found to be associated with an advanced tumor stage. RNA-binding motif 10 overexpression inhibited viability and colony formation capacity of lung adenocarcinoma cell lines and induced cell-cycle arrest at G0/G1 phase in A549 cells and at S phase in H1299 cells. Complementary DNA microarray analysis identified 304 upregulated and 386 downregulated genes induced by RNA-binding motif 10 overexpression, which may be involved in cancer, focal adhesion, peroxisome proliferator-activated receptor–regulated gene pathway, cytokine–cytokine receptor interaction, mitogen-activated protein kinase signaling, complement and coagulation cascades, platelet amyloid precursor protein pathway, extracellular matrix-receptor interaction, and small cell lung cancer–related genes. Expression of FGF2, EGFR, WNT5A, NF-κB, and RAP1A was downregulated, whereas expression of AKT2, BIRC3, and JUN was upregulated. RNA-binding motif 10 messenger RNA and protein were reduced in lung adenocarcinoma tissues, and RNA-binding motif 10 overexpression inhibited lung adenocarcinoma cancer cell malignant behavior in vitro. Molecularly, RNA-binding motif 10 regulates many gene pathways involving in the tumor development or progression.

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