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161.
162.
Mitogen-activated protein (MAP) kinases, a closely related family of protein kinases, are involved in cell cycle regulation
and differentiation in yeast and human cells. They have not been documented in ciliates. We used PCR to amplify DNA sequences
of a ciliated protozoan—Paramecium caudatum—using primers corresponding to amino acid sequences that are common to MAP kinases. We isolated and sequenced one putative
MAP kinase-like serine/threonine kinase cDNA from P. caudatum. This cDNA, called pcstk1 (Paramecium caudatum Serine/Threonine Kinase 1) shared approximately 35% amino acid identity with MAP kinases from yeast. MAP kinases are activated
by phosphorylation of specific threonine and tyrosine residues. These two amino acid residues are conserved in the PCSTK1
sequence at positions Thr 159 and Tyr 161. The PSTAIRE motif, which is characteristic of the CDK2 gene family, cannot be found
in ORF of PCSTK1. The highest homology score was to human STK9, which contains MAP type kinase domains. Comparisons of expression
level have shown that pcstk1 is expressed equally in cells at different stages (sexual and asexual). We discussed the possibility, as in other organisms,
that a family of MAP kinase genes exists in P. caudatum. 相似文献
163.
Molecular evolution of nitrate reductase genes 总被引:9,自引:0,他引:9
To understand the evolutionary mechanisms and relationships of nitrate reductases (NRs), the nucleotide sequences encoding
19 nitrate reductase (NR) genes from 16 species of fungi, algae, and higher plants were analyzed. The NR genes examined show
substantial sequence similarity, particularly within functional domains, and large variations in GC content at the third codon
position and intron number. The intron positions were different between the fungi and plants, but conserved within these groups.
The overall and nonsynonymous substitution rates among fungi, algae, and higher plants were estimated to be 4.33 × 10−10 and 3.29 × 10−10 substitutions per site per year. The three functional domains of NR genes evolved at about one-third of the rate of the N-terminal
and the two hinge regions connecting the functional domains. Relative rate tests suggested that the nonsynonymous substitution
rates were constant among different lineages, while the overall nucleotide substitution rates varied between some lineages.
The phylogenetic trees based on NR genes correspond well with the phylogeny of the organisms determined from systematics and
other molecular studies. Based on the nonsynonymous substitution rate, the divergence time of monocots and dicots was estimated
to be about 340 Myr when the fungi–plant or algae–higher plant divergence times were used as reference points and 191 Myr
when the rice–barley divergence time was used as a reference point. These two estimates are consistent with other estimates
of divergence times based on these reference points. The lack of consistency between these two values appears to be due to
the uncertainty of the reference times.
Received: 10 April 1995 / Accepted: 10 September 1995 相似文献
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165.
构建嵌入第二内含子的甘丙肽(Galanin,GAL)全长基因组cDNA的重构分子。通过RT-PCR扩增出cDNA编区的序列,分别从基因组中扩增出cDNA的5′和3′端部分非编码序列;使用重叠延伸PCR(overlap extention PCR,OE-PCR)方法将三个片段重叠获得全长cDNA序列;再将全长cDNA从第三外显子第15个碱基处分成两部分,分开的cDNA前半部分和后半部分以及第二内含子进行重叠延伸获得重构分子,含有第二内含子的甘丙肽(GAL)全长基因组cDNA;将重构分子连入pMDI9-Tsimple载体。电泳分析观察到清晰的重构分子片段;测序显示重构分子由所设计的序列组成,第二内含子插入的位置准确,且无移码。使用重叠延伸PCR能够成功在cDNA中插入内含子获得一段重构基因。 相似文献
166.
Towards genetic based insect resistance in strawberry using the Cowpea trypsin inhibitor gene 总被引:4,自引:0,他引:4
An efficient shoot regeneration system has been developed involving organo-genesis from stem tissue of the strawberry cultivars Melody, Rhapsody and Symphony. Regeneration also occurred in the presence of antibiotics. The system was shown to be suitable for transformation using the GUS:Intron marker gene construct. Histological analysis using the substrate 5-bromo-4-chloro-3-indolyl B-D-glucuronide (X-Gluc) and the polymerase chain reaction (PCR) using specific primers permitted the identification of transformants.
Inoculations were then carried out using the Cowpea protease trypsin inhibitor (CpTi) gene construct. A simple visual assay technique was developed to identify those plants expressing CpTi, allowing them to be selected for further analysis. Trypsin acts on the substrate α-N-benzoyl-DL-arginine-p-nitroanilide (BAPNA) to produce p-nitroaniline (p-na). Trypsin inhibition CpTi was easily detected as a reduction in or elimination of yellow colour development which occurred as the hydrolysis of BAPNA to pna proceeded (Preiser, Schmitz, Maestracci & Crane, 1975). The polymerase chain reaction (PCR) was used to confirm the presence of the gene sequence. 相似文献
Inoculations were then carried out using the Cowpea protease trypsin inhibitor (CpTi) gene construct. A simple visual assay technique was developed to identify those plants expressing CpTi, allowing them to be selected for further analysis. Trypsin acts on the substrate α-N-benzoyl-DL-arginine-p-nitroanilide (BAPNA) to produce p-nitroaniline (p-na). Trypsin inhibition CpTi was easily detected as a reduction in or elimination of yellow colour development which occurred as the hydrolysis of BAPNA to pna proceeded (Preiser, Schmitz, Maestracci & Crane, 1975). The polymerase chain reaction (PCR) was used to confirm the presence of the gene sequence. 相似文献
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