scholarly journals Cloning and Characterization of a cDNA Encoding Calcium/Calmodulin-Dependent Glutamate Decarboxylase from Scutellaria Baicalensis

2013 ◽  
Vol 8 (9) ◽  
pp. 1934578X1300800
Author(s):  
Yeon Bok Kim ◽  
Md Romij Uddin ◽  
Do Yeon Kwon ◽  
Min-Ki Lee ◽  
Sun-Ju Kim ◽  
...  
Keyword(s):  
Amino Acid ◽  
Glutamate Decarboxylase ◽  
Scutellaria Baicalensis ◽  
Pcr Analysis ◽  
Cdna Clones ◽  
Gamma Aminobutyric Acid ◽  
Accumulation Pattern ◽  
Transcript Levels ◽  
Important Amino Acid ◽  
Gaba Biosynthesis

Gamma-aminobutyric acid (GABA), synthesized by glutamate decarboxylase (GAD), plays an important role in plants. To study the molecular mechanism of GAD regulation and to examine the levels of GABA in Scutellaria baicalensis, we isolated cDNA clones (SbGAD1 and 2) encoding GAD from S. baicalensis. The open reading frames of SbGAD1 and 2 were 1,503 and 1,494 bp long and had 450 and 497 amino acid residues, respectively. Quantitative real-time RT-PCR analysis was performed to show the variation of transcript levels among different organs of S. baicalensis. Transcript levels of SbGAD1 and 2 were highest in the root and flower, respectively. The GABA content of different parts (ranked in descending order) was as follows: leaf > flower > stem > root. We concluded that the expression pattern of SbGAD1 and 2 did not match the accumulation pattern of GABA in different organs. We presume that GABA biosynthesis might be more controlled by SbGAD2 than SbGAD1. These data will aid in future studies that seek to understand the mechanisms underlying GABA biosynthesis, an important amino acid that is synthesized by the GAD enzyme. To explain adequately the GABA biosynthesis mechanisms in S. baicalensis, the enzyme activities of SbGAD1 and 2 should be determined in the near future.

scholarly journals Molecular Cloning and Characterization of Tyrosine Aminotransferase and Hydroxyphenylpyruvate Reductase, and Rosmarinic Acid Accumulation in Scutellaria baicalensis

2014 ◽  
Vol 9 (9) ◽  
pp. 1934578X1400900
Author(s):  
Yeon Bok Kim ◽  
Md Romij Uddin ◽  
YeJi Kim ◽  
Chun Geon Park ◽  
Sang Un Park
Keyword(s):  
Amino Acid ◽  
Rosmarinic Acid ◽  
High Performance ◽  
Molecular Mechanisms ◽  
Tyrosine Aminotransferase ◽  
Open Reading Frames ◽  
Scutellaria Baicalensis ◽  
Cdna Clones ◽  
Amino Acid Residues ◽  
Transcript Levels

Rosmarinic acid (α- O-caffeoyl-3,4-dihydroxyphenylacetic acid, RA) is a caffeoyl ester widely distributed in plants. cDNA clones encoding tyrosine aminotransferase (TAT1 and 2) and hydroxyphenylpyruvate reductase (HPPR) have been isolated from Scutellaria baicalensis. The open reading frames (ORFs) of SbTAT1 and 2 were 1230 and 1272 bp long and encoded 409 and 423 amino acid residues, respectively. HPPR corresponded to a 942-bp ORF and 313 amino acid residues of translated protein. To study the molecular mechanisms of TAT and HPPR and investigate RA accumulation in S. baicalensis, we examined the transcript levels of TAT isoforms and HPPR with quantitative real-time PCR and analyzed the RA content in different organs by using high-performance liquid chromatography. The transcript levels of SbTAT1 SbTAT2, and SbHPPR in the flowers were higher than those in other organs. RA was also highly accumulated in the flowers and with a trace amount in the roots. No RA was detected in the leaves and stems of S. baicalensis. The amount of accumulated RA in the flowers was 28.7 times higher than that in the roots. Our results will be helpful in elucidating the mechanisms of RA biosynthesis in S. baicalensis.

Cloning of rabbit prolactin cDNA and prolactin gene expression in the rabbit mammary gland

1996 ◽  
Vol 16 (1) ◽  
pp. 27-37 ◽  
Author(s):  
L Gabou ◽  
M Boisnard ◽  
I Gourdou ◽  
H Jammes ◽  
J-P Dulor ◽  
...  
Keyword(s):  
Gene Expression ◽  
Mammary Gland ◽  
Amino Acid ◽  
Untranslated Regions ◽  
Pcr Analysis ◽  
Cdna Clones ◽  
Rt Pcr ◽  
Prolactin Gene ◽  
New Zealand White Rabbits ◽  
Rna Probe

ABSTRACT cDNA clones coding for rabbit prolactin were isolated from a pituitary library using a rat prolactin RNA probe. One cDNA contained 873 bases including the entire coding sequence of rabbit prolactin, its signal peptide and the 5′ and 3′ untranslated regions of 44 and 145 nucleotides respectively. The deduced amino acid sequence of the cloned prolactin cDNA presented a 93–78% identity with mink, porcine and human prolactins. The prolactin gene transcription was investigated by RT-PCR analysis in several organs of midlactating New Zealand White rabbits. The ectopic transcription of the prolactin gene was examined in more detail in the mammary gland. A strong PCR signal was detected in the mammary gland of virgin does and was also observed during pregnancy and at the beginning of lactation. This PCR signal was very weak in mid-lactating and absent in post-weaning mammary gland.

scholarly journals Identification and tissue distribution of mRNAs encoding salmon-type calcitonins-IV and -V in the rainbow trout

2004 ◽  
Vol 32 (3) ◽  
pp. 963-974 ◽  
Author(s):  
Y Hidaka ◽  
M Suzuki
Keyword(s):  
Rainbow Trout ◽  
Amino Acid ◽  
Tissue Distribution ◽  
Restriction Enzymes ◽  
Tissue Expression ◽  
Pcr Analysis ◽  
Cdna Clones ◽  
Rt Pcr ◽  
Parenchymal Cells ◽  
Ultimobranchial Gland

Four types of calcitonin are produced in salmonid fish, although their functional diversity is almost unknown. To explore the significance of these isoforms, we have characterized salmon-type calcitonin (sCT) mRNAs in the rainbow trout (Oncorhynchus mykiss), and examined their tissue distribution. In addition to the previously isolated sCT-I cDNAs, two new forms of sCT cDNA were cloned from the ultimobranchial gland, and one of them (sCT-IV cDNA) was predicted to encode an N-terminal peptide of 80 amino acid residues, a putative cleavage site Lys-Arg, sCT-IV, a cleavage and amidation sequence Gly-Lys-Lys-Arg, and a C-terminal peptide of 18 amino acids. The sCT-IV precursor was 78% identical with the rainbow trout sCT-I precursors. The other cloned cDNA encoded a precursor for a novel CT, sCT-V. The sCT-V peptide was different from sCT-IV by only one amino acid residue: Val at position 8 in the latter was replaced by Met. The sCT-V precursor had 80 and 90% identity with the sCT-I and -IV precursors respectively. No cDNA clones were obtained for sCTs-II or -III.Tissue distribution of sCT-I, -IV and -V mRNAs was examined by RT-PCR and specific cleavage with restriction enzymes. An amplified fragment from sCT-I mRNA was detected not only in the ultimobranchial gland, but also in the gills, testis and ovary. RT-PCR analysis coupled to restriction digestion further revealed that sCT-IV mRNA was expressed in both the testis and the ultimobranchial gland. The expression sites of sCT-IV mRNA were localized to the Leydig cells of the testis and to the parenchymal cells of the ultimobranchial gland, by in situ hybridization histochemistry. Although the amino acid sequence of sCT-V peptide was nearly the same as that of sCT-IV, the sCT-V gene showed a much wider pattern of expression: the band amplified by RT-PCR was detected in all the tissues examined except the kidney, gills and blood cells. The sCT-V mRNA was shown to be localized in the parenchymal cells of the ultimobranchial gland, but not in other tissues at the cellular level, suggesting very low expression of sCT-V mRNA in those tissues. Our results show different patterns of tissue expression of three types of sCT genes in the rainbow trout, suggesting that sCTs-I, -IV and -V might differ in their local actions.

Structures and expression patterns of two cDNA clones encoding S-adenosyl-L-methionine synthetase from the root nodule of Elaeagnus umbellata

2001 ◽  
Vol 28 (9) ◽  
pp. 951
Author(s):  
Sang Ho Lee ◽  
Ho Bang Kim ◽  
Chung Sun An
Keyword(s):  
Amino Acid ◽  
Expression Pattern ◽  
Root Nodule ◽  
Vascular System ◽  
Expression Patterns ◽  
Amino Acid Level ◽  
Nodule Development ◽  
Pcr Analysis ◽  
Cdna Clones ◽  
Elaeagnus Umbellata

This paper originates from an address at the 8th International Symposium on Nitrogen Fixation with Non-Legumes, Sydney, NSW, December 2000 Two cDNA clones encoding S-adenosyl-L-methionine synthetase (SAMS) were isolated from the root nodule cDNA library of Elaeagnus umbellata Thunberg and analysed on the basis of deduced amino acid sequence and expression pattern. Two EuSAMS clones shared 75–84% identity at the nucleotide level, and 85–95% identity at the amino acid level, with the other plant SAMS genes. Genomic Southern hybridization revealed the presence of more than two copies of SAMS genes in the genome of E. umbellata. Reverse transcriptase-mediated polymerase chain reaction (RT–PCR) analysis showed EuSAMS1 transcripts were more abundant than those of EuSAMS2. Similar to the expression pattern of other plant SAMS genes, both genes were expressed at higher levels in root than in leaf. During nodule development, expression of both genes was increased, with the highest level at 6–8 week after inoculation, and decreased rapidly thereafter. In situ hybridization analysis also showed both SAMS transcripts in the meristem zone, the infected cells of the fixation zone and in the central vascular system of root nodules. However, EuSAMS2 transcripts were strongly detected in the prefixation zone, whereas EuSAMS1 transcripts were hardly detected. These results suggest different regulatory mechanisms for the two genes in the root nodule. The expression pattern of SAMS genes in the root nodule may correlate mostly with cell wall synthesis, polyamine biosynthesis and other methylation-mediated functions.

scholarly journals 263 IDENTIFICATION OF A NOVEL MOPT GENE IN HUMAN AND MOUSE ADULT TESTIS

2005 ◽  
Vol 17 (2) ◽  
pp. 281
Author(s):  
Y.-J. Choi ◽  
S.-J. Kang ◽  
J.-H. Kim
Keyword(s):  
Amino Acid ◽  
Genomic Dna ◽  
Wild Type Mouse ◽  
Amino Acid Sequences ◽  
Mouse Tissue ◽  
Pcr Analysis ◽  
Human Testis ◽  
Testicular Development ◽  
Cdna Clones ◽  
Late Spermatid

To discover late stage germ cell-specific transcripts we prepared a cDNA library from adult testes of 35-day old mice and subtracted it with mRNA from the testes of juvenile mice. Real-time RT-PCR analysis indicated that 42 cDNA clones in the subtracted library were expressed more intensely in the adult testes than in the juvenile testes. One clone identified by subtraction is expressed preferentially in the late spermatid and is located on chromosome 17E3 in mouse and 2p22 in human. The full nucleotide and amino acid sequences of mouse and human MOPT gene are deposited in EMBL GenBank (AY367765 And AY367766). Human MOPT is spliced by 5 exons and 4 introns and encompasses 7,000 bp of genomic DNA (from bp 355 822 to 425 511) of NT-022184.13, whereas mouse MOPT is spliced by 5 exons and 4 introns and encompasses 7,382 bp of genomic DNA (from bp 6227407 to 6235588) of NT-039658.2. Because of the limited availability of human testis samples, development-dependent expression of MOPT mRNA was conducted using its mouse homologue and semiquantitative PCR. The number of cycles completed before entering the exponential growth, recorded by amplifier PE5700 for mouse MOPT, were 1.11 ± 0.23, 1.05 ± 0.04, 1.5 ± 0.2, 5.55 ± 0.65, 19.35 ± 0.65, 68.65 ± 2.15, and 185.15 ± 6.15 in W/W, postnatal day 5-, 8-, 12-, 15-, 18-, 22-, and 28-day mouse tissue samples, respectively. The difference among the three times was significant (P < 0.01, ANOVA). These results suggest that expression of MOPT gene increased from postnatal Day 5 to Day 28, indicating possible involvement in testicular development. The ORF encodes a protein containing 79 amino acid residues. A MORN motif, EGQFKDNMFHGLGTYTFPNG, was identified in the predicted protein sequence of MOPT; function of this motif is unknown. In situ hybridization of 12-week-old wild-type mouse testes using an antisense riboprobe and immuno-gold data indicated MOPT was expressed as a late spermatid and acrosome reaction. This work was supported by BK21 program.

Purification of Antihemophilic Factor (Factor VIII) by Amino Acid Precipitation*

1964 ◽  
Vol 11 (01) ◽  
pp. 064-074 ◽  
Author(s):  
Robert H Wagner ◽  
William D McLester ◽  
Marion Smith ◽  
K. M Brinkhous
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Factor Viii ◽  
Plasma Protein ◽  
Acid Precipitation ◽  
Aminobutyric Acid ◽  
Gamma Aminobutyric Acid ◽  
Specific Procedure ◽  
Beta Alanine ◽  
Antihemophilic Factor

Summary1. The use of several amino acids, glycine, alpha-aminobutyric acid, alanine, beta-alanine, and gamma-aminobutyric acid, as plasma protein precipitants is described.2. A specific procedure is detailed for the preparation of canine antihemophilic factor (AHF, Factor VIII) in which glycine, beta-alanine, and gammaaminobutyric acid serve as the protein precipitants.3. Preliminary results are reported for the precipitation of bovine and human AHF with amino acids.

scholarly journals Cloning and sequence analysis of cDNAs encoding the α1, α2 and α3 chains of mouse collagen VI

1993 ◽  
Vol 291 (3) ◽  
pp. 787-792 ◽  
Author(s):  
R Z Zhang ◽  
T C Pan ◽  
R Timpl ◽  
M L Chu
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequences ◽  
Untranslated Regions ◽  
Cdna Libraries ◽  
Cdna Clones ◽  
Collagen Vi ◽  
Central Segment ◽  
Glycosylation Sites ◽  
Key Features ◽  
Triple Helical Domain

cDNA clones encoding the alpha 1, alpha 2 and alpha 3 chains of mouse collagen VI have been isolated by screening cDNA libraries with the corresponding human probes. The composite cDNAs for the alpha 1, alpha 2, and alpha 3 chains are 2.5, 1.6 and 2.9 kb in size respectively. The alpha 1 and alpha 2 cDNAs encode the C-terminal portions of the chains as well as the entire 3′-untranslated regions, while the alpha 3 cDNAs encode a central segment of 959 amino acids flanking the triple-helical domain. The deduced amino acid sequences share 86-88% identity with the human counterparts and 67-73% identity with the chicken equivalents. Alignment of the deduced amino acid sequences of mouse, human and chicken collagens reveal that the key features of the protein, including the cysteine residues, imperfections in the Gly-Xaa-Xaa regions, Arg-Gly-Asp sequences and potential N-glycosylation sites, are mostly conserved.

Important Amino Acid Residues that Confer CYP2C19 Selective Activity to CYP2C9

2008 ◽  
Vol 144 (3) ◽  
pp. 323-333 ◽  
Author(s):  
Y. Wada ◽  
M. Mitsuda ◽  
Y. Ishihara ◽  
M. Watanabe ◽  
M. Iwasaki ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Residues ◽  
Important Amino Acid ◽  
Selective Activity
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