Amino acids and peptides. LVII. Synthetic peptide with a sequence of ribonuclease from Sulfolobus solfataricus, SSR(1-62), does not function as an RNase

The 62 residue peptide, SSR(1-62), whose sequence corresponds to that of ribonuclease (RNase) from Sulfolobus solfataricus, and its related peptides, SSR(1-22) and SSR(10-62), were chemically synthesized and their RNase activity and DNA-binding activity were examined. The RNase activity assay using yeast RNA or tRNA(fMet) as substrate showed that the synthetic peptide SSR(1-62) did not hydrolyze yeast RNA or tRNA(fMet). These data were not consistent with previous reports that both the native peptide isolated from S. solfataricus [Fusi et al. (1993) Eur. J. Biochem. 211, 305-311] and the recombinant peptide expressed in Escherichia coli [Fusi et al. (1995) Gene 154, 99-103] were able to hydrolyze tRNA(fMet). However, the synthetic SSR(1-62) exhibited DNA-binding activity. In the presence of synthetic SSR(1-62), the cleavage of DNA (plasmid pUCRh2-4) by restriction endonuclease (EcoRI) was not observed, suggesting that synthetic SSR(1-62) bound to DNA protected DNA from its enzymatic digestion. Neither SSR(1-22) nor SSR(10-62) prevented DNA from being cleaved by a restriction enzyme. These findings strongly suggest the importance of not only the N-terminal region of SSR(1-62) but also the C-terminal region for DNA-binding. Circular dichroism spectroscopy of synthetic SSR(1-62) indicated a beta-sheet conformation, in contrast with synthetic SSR(1-22), which exhibited an unordered conformation.     

Secondary sex ratio variation in the Cayo Santiago macaque population

Secondary sex ratios (SSR) were calculated from 1,385 offspring delivered by 372 females in the Cayo Santiago population of free-ranging rhesus monkeys (Macaca mulatta) from 1976 through 1984. The SSR for the entire colony ranged from 0.86 to 1.46 males per female (combined total: 1.08), but no significant difference was observed (P > .05). SSR values were compared among the troops for each year. The SSR differed significantly among the six social groups (P < .05) only in 1978. The annual SSR of each troop was compared over 9 years. Significant variation was found only in group O. The annual SSR was significantly skewed (P < .05, males > females) for three troops in 3 separate years. The SSR did not vary according to troop rank. No significant difference was found among the 17 matrilines of the population, but comparison of matrilines within each social group revealed a significant difference in the SSR (P < .02) for the three matrilines in group I. This was due to the significantly skewed SSR (P = .0080, females > males) of the DM genealogy in that troop. SSR values were not related to matrilineal rank. Individual dominance rank did not bias the SSR. Complete reproductive histories for 266 females showed no evidence of significantly skewed SSR values. Age-related effects on the SSR were examined by using cross-sectional and cohort-based analyses. The SSR did not vary significantly (P > .05) with maternal age, but it was significantly skewed (P < .05) toward males at the ages of 5 and 9 years. Parity had no significant effect (P > .05) on SSR values. Wide variation occurred in the SSR of the Cayo Santiago population. Rank-related adjustment of the SSR at the level of the troop, matriline, or individual, as reported in short-term studies of other primate social groups, may reflect normal annual variation in the SSR evident only from longitudinal observations of large multigroup primate populations.     

大白菜相对于结球甘蓝连锁群特异SSR标记的筛选

以9个大白菜品种和10个结球甘蓝品种为试材,对位于芸薹属A基因组10个连锁群的207对SSR引物进行PCR筛选,共筛选出33个大白菜相对于结球甘蓝的特异SSR标记,涉及了大白菜的10个连锁群,其中位于A1连锁群3对,A2连锁群4对,A3连锁群5对,A4连锁群2对,A5连锁群3对,A6连锁群4对,A7连锁群2对,A8连锁群1对,A9连锁群2对,A10连锁群7对。为进一步利用SSR标记鉴定结球甘蓝—大白菜异附加系奠定了基础。     

An improved technique for isolating codominant compound microsatellite markers

An approach for developing codominant polymorphic markers (compound microsatellite (SSR) markers), with substantial time and cost savings, is introduced in this paper. In this technique, fragments flanked by a compound SSR sequence at one end were amplified from the constructed DNA library using compound SSR primer (AC)₆(AG)₅ or (TC)₆(AC)₅ and an adaptor primer for the suppression-PCR. A locus-specific primer was designed from the sequence flanking the compound SSR. The primer pairs of the locus-specific and compound SSR primers were used as a compound SSR marker. Because only one locus-specific primer was needed for design of each marker and only a common compound SSR primer was needed as the fluorescence-labeled primer for analyzing all the compound SSR markers, this approach substantially reduced the cost of developing codominant markers and analyzing their polymorphism. We have demonstrated this technique for Dendropanax trifidus and easily developed 11 codominant markers with high polymorphism for D. trifidus. Use of the technique for successful isolation of codominant compound SSR markers for several other plant species is currently in progress.     

Large-scale development of PIP and SSR markers and their complementary applied in Nicotiana

PIP (Potential Intron Polymorphism) and SSR (Simple Sequence Repeats) were used in many species, but large-scale development and combined use of these two markers have not been reported in tobacco. In this study, a total of 12,388 PIP and 76,848 SSR markers were designed and uploaded to a webaccessible database (http://yancao.sdau.edu.cn/tgb/). E-PCR analysis showed that PIP and SSR rarely over-lapped and were strongly complementary in the tobacco genome. The density of markers was 3.07 PIP and 1.72 SSR per 10 kb of the known sequences. A total of 153 and 166 alleles were detected by 22 PIP and 22 SSR markers in 64 Nicotiana accessions. SSR produced higher PIC (polymorphism information content) values and identified more alleles than PIP, whereas PIP could identify larger numbers of rare alleles. Mantel testing demonstrated a high correlation coefficient (r = 0.949, P < 0.001) between PIP and SSR. The UPGMA dendrogram created from the combined PIP and SSR markers was clearer and more reliable than the individual PIP or SSR dendrograms. It suggested that PIP and SSR can make up the deficiency of molecular markers not only in tobacco but other plant.     

SSR repeat dynamics in mitochondrial genomes of five domestic animal species

SSR (simple sequence repeats) are ubiquitously abundant in genomes. In organellar mitochondrial genome of animals, its distribution, size dynamics and effectiveness for phylogenetic relationship have not been understood. Present investigation reveals organisation of SSR in genic and intergenic region, its length and repeat motif dynamics, extent of conservation of flanking regions, appropriateness of these SSR data in establishing phylogenetic relationship. Contrary to eukaryotic nuclear abundance of SSR in non-coding region, we found abundance in coding region. Like nuclear SSR, most hyper mutable repeats were found in non coding region having di nucleotide motifs of mitochondrial genome but contrary to human having high mutable tetra repeats in case of mitochondrial genomes this was found to be with tri-motif repeats. SSR of mitochondrial genomes also show cyclical expansion and shrinkage in pattern of SHM (simple harmonic motion) with respect to time its non- linear thus not appropriate for phylogenetic analysis though the flanking regions of these SSR also conserved like nuclear SSR.     

基于SSR标记的贵州薏苡种质资源遗传多样性?分析

利用SSR标记研究了22份薏苡种质的遗传多样性,用11对扩增带型稳定的SSR引物从供试材料中检测出105个等位基因变异,每对引物检测等位基因4～20个,平均9.55个。SSR引物的PIC介于0.3048～0.9238,平均多态性信息量为0.8255。利用UPGMA聚类分系法将供试自交系划分为4类,该划分结果与根据地理来源、种质系谱的分类结果基本一致。SSR分子标记辅助的种质改良是薏苡品种改良的重要途径。     

Identification and development of polymorphic EST-SSR markers by sequence alignment in pepper, Capsicum annuum (Solanaceae)

? Premise of the study: The redundancies in expressed sequence tags (ESTs) in the National Center for Biotechnology Information sequence database were used to identify and develop polymorphic simple sequence repeat (SSR) markers for pepper (Capsicum annuum). ? Methods and Results: Sixty-eight polymorphic SSR loci were identified in the contigs (containing redundant ESTs) generated by assembling 118060 pepper ESTs from the public sequence database. Thirty-three SSR markers exhibited polymorphism among 31 pepper varieties, with alleles per SSR marker ranging from two to six. The mean observed and expected heterozygosity were 0.28 and 0.39, respectively. There were 18 SSR markers with a motif repeat number of less than five, accounting for 55% of the total. ? Conclusions: We demonstrated the value of mining the redundant sequences in public sequence databases for the development of polymorphic SSR markers, which can be used for marker-assisted breeding in pepper.     

Genetic diversity among Japanese indigenous common buckwheat (Fagopyrum esculentum) cultivars as determined from amplified fragment length polymorphism and simple sequence repeat markers and quantitative agronomic traits.

We assessed the genetic diversity in Japanese indigenous common buckwheat (Fagopyrum esculentum) cultivars using amplified fragment length polymorphism (AFLP) and simple sequence repeat (SSR) markers and investigated the relationships between the genetic diversity and agronomic traits. The average expected intracultivar hetero zygosity was 0.303 for AFLP and 0.819 for SSR. The differentiations among agroecotypes, among cultivars within an agroecotype, and among cultivars were small (0.002, 0.024, and 0.026 for SSR and 0.013, 0.013, and 0.026 for AFLP, respectively) but statistically significant from zero except for the SSR differentiation among agroecotypes. In principal coordinates analysis, cultivars within the same agroecotype tended to cluster, indicating that agroecotypes well reflected the genetic relationships among cultivars. In AFLP, the differentiation among the agroecotypes was more distinct than in SSR, and genetic distance showed a moderate correlation with the difference in quantitative traits, indicating that AFLP can resolve the relationships among cultivars with better resolution than SSR. By contrast, SSR may be more sensitive to demographic changes. Four of the five SSR markers showed a significant positive correlation (Kendall's tau = 0.382-0.607) between allelic richness and variation in flowering timing, indicating that cumulative bottleneck events have occurred during the population history, with a decline in the variation of photosensitivity of flowering.     

PCR-multiplexes for a genome-wide framework of simple sequence repeat marker loci in cultivated sunflower

Simple sequence repeat (SSR) and other DNA sequence-tagged site markers can be genotyped more rapidly and cost efficiently by simultaneously amplifying multiple loci (multiplex PCR). The development of PCR-multiplexes for a nearly genome-wide framework of 78 SSR marker loci in cultivated sunflower ( Helianthus annuus L.) is described herein. The most outstanding single-locus SSR markers in the public collection (300 out of 1,089) were identified and screened for polymorphisms among 24 elite inbred lines, preparatory to selecting SSR markers for testing in multiplex PCRs. The selected SSR markers produced robust PCR products, amplified a single locus each, were polymorphic among elite inbred lines (minimum, mean and maximum heterozygosities were 0.08, 0.53 and 0.85, respectively), and supply a dense genome-wide framework of predominantly or completely codominant, single-locus DNA markers for molecular breeding and genomics research in sunflower. Thirteen six-locus multiplex PCRs were developed for 78 SSR marker loci strategically positioned throughout the sunflower genome (three to five per linkage group) by identifying compatible SSR primer combinations and optimizing multiplex PCR protocols. The multiplexed SSR markers, when coupled with 17 complementary SSR marker loci, create a 'standard genotyping' set ideal for first-pass scans of the genome, as are often needed when screening bulked-segregant DNA samples or mapping phenotypic trait loci. The minimum, mean and maximum heterozygosities of the multiplexed SSR markers were 0.38, 0.62 and 0.83, respectively. The PCR-multiplexes increase genotyping throughput, reduce reagent costs, and are ideal for repetitive genotyping applications where common sets of SSR marker loci are required or advantageous.     

花生微卫星标记的研究进展

近年来花生微卫星标记的开发取得了一定的进展, 初步揭示了花生在DNA水平上的遗传多样性。花生微卫星标记的开发途径主要包括通过构建小片段基因组文库开发基因组SSR标记, 根据花生EST序列开发EST-SSR标记, 根据豆科植物序 列信息和SSR标记开发花生SSR标记, 将SSR标记与其它分子标记结合开发新的DNA标记, 以及基于SSR核心序列开发ISSR标记。花生微卫星标记主要应用于遗传多样性研究、遗传图谱与品种指纹图谱构建以及分子标记辅助育种等领域。本文综述了花生SSR标记开发研究的进展及应用。     

编码区和非编码区SSR标记对水稻类群的比较研究

设计14对水稻编码区SSR引物和选取已公布的非编码区SSR引物12对、编码区SSR引物3对,采用SSR技术,对29个标记在60个水稻材料中的多态性进行分析。结果表明,编码区SSR标记平均检测到3.59个多态性位点,多态信息量PIC(polymorphism information conten)在0.032～P0.853之间,平均值为0.447;非编码区SSR标记平均检测到3.92个多态性位点,PIC在0.063～P0.795之间,平均值为0.521。聚类分析显示,非编码区SSR标记能更加精确地区分来自不同地区的水稻类群,编码区SSR标记也具有良好的多态性,同样可以用于分析水稻的亲缘关系。     

Overexpression of SSR2 promotes proliferation of liver cancer cells and predicts prognosis of patients with hepatocellular carcinoma

Signal Sequence Receptor Subunit 2 (SSR2) is a key endoplasmic reticulum gene involved in protein folding and processing. Previous studies found that it was upregulated in several cancers, but its precise role in hepatocellular carcinoma (HCC) remains unclear. To have a better understanding of this gene in HCC, we examined the expression of SSR2 in HCC tissues by analysing The Cancer Genome Atlas (TCGA) data and immunohistochemistry. We also assessed the association between SSR2 expression and clinicopathological characteristics of HCC patients and patient survival. Potential function of SSR2 was predicted through GSEA and protein–protein interaction analysis. MTT, flowcytometry, transwell and a nude mice xenograft model were employed to investigate the biological functions in vivo and in vitro. The results showed that the expression of SSR2 was significantly increased in HCC tissues, and SSR2 expression was associated with several clinical characteristics. In addition, patients with higher SSR2 expression had poorer survival. Enrichment analysis suggested that SSR2 was probably involved in biological process or signalling pathways related to G2/M checkpoint, passive transmembrane transporter activity, ATF2_S_UP. V1_UP and ncRNA metabolic process. Further experimental study showed that SSR2 knockdown inhibited cell proliferation, migration and invasion ability and promoted apoptosis and cell cycle arrest in vitro. Moreover, downregulation of SSR2 also repressed the growth of HepG2 cells in vivo. In conclusion, our study suggests that SSR2 may act as an oncogene in HCC.     

Comparative Assessment of SSR and AFLP Markers for Evaluation of Genetic Diversity and Conservation of Fig, <Emphasis Type="Italic">Ficus carica</Emphasis> L., Genetic Resources in Tunisia

This study characterises the genetic variability of fig, Ficus carica L., using simple sequence repeat (SSR) and amplified fragment length polymorphism (AFLP) markers. It compares the efficiency and utility of the two techniques in detecting variation and establishing genetic relationships among Tunisian fig cultivars. Our results show that using both marker systems, the Tunisian fig germ plasm is characterised by having a large genetic diversity at the deoxyribonucleic acid level, as most of AFLP bands were detected and all SSR markers were polymorphic. In fact, 351 (342 polymorphic) and 57 (57 polymorphic) bands were detected using AFLP and SSR primers, respectively. SSR markers were the most polymorphic with an average polymorphic information content value of 0.94, while AFLP markers showed the highest effective multiplex ratio (56.9) and marker index (45.2). The effective marker index was recorded highest (4.19) for AFLP markers and lowest (0.70) for the SSR ones. Our results demonstrate that (1) independent as well as combined analyses of cluster analyses of SSR and AFLP fragments showed that cultivars are clustered independently from their geographical origin, horticultural classifications and tree sex; (2) the analysis of molecular variance allowed the partitioning of genetic variation within and among fig groups and showed greater variation within groups and (3) AFLP and SSR markers datasets showed positive correlation. This study suggests the SSR and AFLP markers are suitable for diversity analysis and cultivars fingerprinting. An understanding of the genetic diversity and population structure of F. carica in Tunisia can also provide insight into the conservation and management of this species.     

Genetic effects on microsatellite diversity in wild emmer wheat (Triticum dicoccoides) at the Yehudiyya microsite,Israel

This study investigated allele size constraints and clustering, and genetic effects on microsatellite (simple sequence repeat, SSR) diversity at 28 loci comprising seven types of tandem repeated dinucleotide motifs in a natural population of wild emmer wheat, Triticum dicoccoides, from a shade vs sun microsite in Yehudiyya, northeast of the Sea of Galilee, Israel. It was found that allele distribution at SSR loci is clustered and constrained with lower or higher boundary. This may imply that SSR have functional significance and natural constraints. Genetic factors, involving genome, chromosome, motif, and locus significantly affected SSR diversity. Genome B appeared to have a larger average repeat number (ARN), but lower variance in repeat number (sigma(ARN)(2)), and smaller number of alleles per locus than genome A. SSRs with compound motifs showed larger ARN than those with perfect motifs. The effects of replication slippage and recombinational effects (eg, unequal crossing over) on SSR diversity varied with SSR motifs. Ecological stresses (sun vs shade) may affect mutational mechanisms, influencing the level of SSR diversity by both processes.     

Development and mapping of a public reference set of SSR markers in Lolium perenne L.

We report on the characterization and mapping of 76 simple sequence repeat (SSR) markers for Lolium perenne. These markers are publicly available or obtained either from genomic libraries enriched for SSR motifs or L. perenne expressed sequence tag (EST) clones. Four L. perenne mapping populations were used to map the SSR markers. A consensus linkage map of the four mapping populations containing 65 of the SSR markers is presented, together with primer information and a quality score indicating the usefulness of the SSR marker in different populations. The SSR markers identified all seven L. perenne linkage groups.     

Development and use of anchored‐SSRs to study DNA polymorphism in bread wheat (Triticum aestivum L.)

In bread wheat, 21 anchored simple sequence repeat (SSR) primer pairs detecting SSR length polymorphism and 42 anchored SSR primers detecting microsatellite‐anchored fragment length polymorphisms (MFLPs) are reported. Eight bread wheat genotypes were used for detecting polymorphism. The number of alleles in SSR analysis ranged from two to six, with a mean of 2.9 alleles per SSR. The number of polymorphic bands in MFLP ranged from two to 40, with a mean of 12.74 polymorphic bands/primer combination, the SSRs with CT/GA motifs giving the highest level of polymorphism (a mean of 18.37 bands). The average value of polymorphic information content (PIC) was 0.473 for SSRs and 0.061 for MFLP.     

An integrated SSR and RFLP linkage map of Sorghum bicolor (L.) Moench.

We report the development, testing, and use (for genetic mapping) of a large number of polymerase chain reaction (PCR) primer sets that amplify DNA simple sequence repeat (SSR) loci of Sorghum bicolor (L.) Moench. Most of the primer sets were developed from clones isolated from two sorghum bacterial artificial chromosome (BAC) libraries and three enriched sorghum genomic-DNA (gDNA) libraries. A few were developed from sorghum DNA sequences present in public databases. The libraries were probed with radiolabeled di- and trinucleotide oligomers, the BAC libraries with four and six oligomers, respectively, and the enriched gDNA libraries with four and three oligomers, respectively. Both types of libraries were markedly enriched for SSRs relative to a size-fractionated gDNA library studied earlier. However, only 2% of the sequenced clones obtained from the size-fractionated gDNA library lacked a SSR, whereas 13% and 17% of the sequenced clones obtained from the BAC and enriched gDNA libraries, respectively, lacked a SSR. Primer sets were produced for 313 SSR loci. Two-hundred sixty-six (85%) of the loci were amplified and 165 (53%) of the loci were found to be polymorphic in a population composed of 18 diverse sorghum lines. (AG/TC)n and (AC/TG)n repeats comprised 91% of the dinucleotide SSRs and 52% of all of the SSRs at the polymorphic loci, whereas four types of repeats comprised 66% of the trinucleotide SSRs at the loci. Primer sequences are reported for the 165 polymorphic loci and for eight monomorphic loci that have a high degree of homology to genes. Also reported are the genetic map locations of 113 novel SSR loci (including four SSR-containing gene loci) and a linkage map composed of 147 SSR loci and 323 RFLP (restriction fragment length polymorphism) loci. The number of SSR loci per linkage group ranges from 8 to 30. The SSR loci are distributed relatively evenly throughout approximately 75% of the 1406-cM linkage map, but segments of five linkage groups comprising about 25% of the map either lack or contain few SSR loci. Mapping of SSR loci isolated from BAC clones located to these segments is likely to be the most efficient method for placing SSR loci in the segments.     

Influences of experimental factors on spinal stretch reflex latency and amplitude in the human triceps surae.

The spinal stretch reflex (SSR) is commonly assessed via electromyographic (EMG) analysis of joint perturbations inducing changes in muscle length. Previous literature indicates that when large experimental changes in magnitude of agonist background EMG, perturbation velocity, and perturbation amplitude are employed, SSR latency and amplitude are significantly altered. The purpose of this investigation was to evaluate the relative dependence of SSR latency and amplitude on inherent variability in these experimental variables. Soleus SSR latency and amplitude were assessed in 40 healthy subjects following dorsiflexion perturbation under an active state ( approximately 14% MVC). Experimental variables displayed limited variability (means +/- SD): soleus background EMG (13.47 +/- 7.08% MVC), perturbation velocity (96.1 +/- 30 degrees /s), and perturbation amplitude (4 +/- 1 degrees ). SSR latency was not significantly related to soleus background EMG (r = 0.189), perturbation velocity (r = 0.213), or perturbation amplitude (r = 0.202). Similarly, SSR amplitude was not significantly related to soleus background EMG (r = 0.306), perturbation velocity (r = 0.053), or perturbation amplitude (r = 0.056). Variability in experimental variables was much smaller than what has been reported in the literature to significantly impact SSR characteristics. These results suggest that SSR latency and amplitude are independent of agonist background EMG, perturbation velocity, and perturbation amplitude when experimental variability is relatively limited.