Study of molecular genetic diversity and evolutionary history of medicinally important endangered genus Chlorophytum using DNA barcodes

This study was aimed to authenticate and present phylogenetic relationship among 19 species of genus Chlorophytum using DNA barcoding. In all, 107 accessions were analyzed with eight plastid (matK, rbcL, trnH-psbA, rpoC1, ycf5, rpoB, atp and psbK-psbI) and six nuclear (ITS) markers. The matK and rbcL were found to be ideal markers for identification and discrimination of Chlorophytum species. Phylogenetic analysis based on matK and rbcL sequences resolved the species in two major clades. All markers, except matK and rbcL, showed ambiguous reads and paralogy in analysis. DGGE analysis showed the presence of pseudogenes and/or co-amplification in these markers, which caused poor sequence quality. Phylogeny and probable evolution of genus Chlorophytum was proposed on the basis of cytological, morphological and genetic information.     

水稻抗性基因定位及相关分子标记研究进展

水稻是一种重要的粮食作物。而选育高抗性良种是有效防治病虫的危害,增加水稻单位面积产量的一项关键措施。了解水稻本身抗性的遗传信息是进行抗性育种的基础。现代生物技术的发展为抗性育种提供了新途径。本文较系统地概述了水稻对稻瘟病、白叶枯病、稻飞虱、稻叶蝉抗性基因定位及相关分子标记研究的最新发展,为利用分子标记进行水稻抗性育种及抗性基因克隆提供参考文献。     

Association analysis for agronomic traits in wheat under terminal heat stress

Terminal heat stress causes irreversible damage to wheat crop productivity. It reduces the vegetative growth and flowering period that consequently declines the efficiency to capture available stem reserves (carbohydrates) in grains. Markers associated with thermotolerant traits ease in marker assisted selection (MAS) for crop improvement. It identifies the genomic regions associated with thermotolerant traits in wheat, but the scarcity of markers is the major hindrance in crop improvement. Therefore, 158 wheat genotypes were subjected to genotyping with 165 simple sequence repeat markers dispersed on three genomes (A, B and D). Allelic frequency and polymorphic information content values were highest on genome A (5.34 (14% greater than the lowest value at genome D) and 0.715 (3% greater than the lowest value at genome D)), chromosome 4 (5.40 (16% greater than the lowest value at chromosome 2) and 0.725 (5% greater than the lowest value at chromosome 6)) and marker xgwm44 (13.0 (84% greater than the lowest value at marker xbarc148) and 0.916 (46% greater than the lowest value at marker xbarc148)). Bayesian based population structure discriminated the wheat genotypes into seven groups based on genetic similarity indicating their ancestral origin and geographical ecotype. Linkage disequilibrium pattern had highest significant (P < 0.001) linked loci pairs 732 on genome A at r² > 0.1 whereas, 58 on genome B at r² > 0.5. Linkage disequilibrium decay (P < 0.01 and r² > 0.1) had larger LD block (5–10 cM) on genome A. Highly significant MTAs (P < 0.000061) under heat stress conditions were identified for flag leaf area (xwmc336), spikelet per spike (xwmc553), grains per spike (cxfa2147, xwmc418 and xwmc121), biomass (xbarc7) and grain yield (xcfa2147 and xwmc671). The identified markers in this study could facilitate in MAS and gene pyramiding against heat stress in wheat.     

Accuracy map of an optical motion capture system with 42 or 21 cameras in a large measurement volume

Optical motion capture is commonly used in biomechanics to measure human kinematics. However, no studies have yet examined the accuracy of optical motion capture in a large capture volume (>100 m³), or how accuracy varies from the center to the extreme edges of the capture volume. This study measured the dynamic 3D errors of an optical motion capture system composed of 42 OptiTrack Prime 41 cameras (capture volume of 135 m³) by comparing the motion of a single marker to the motion reported by a ThorLabs linear motion stage. After spline interpolating the data, it was found that 97% of the capture area had error below 200 μm. When the same analysis was performed using only half (21) of the cameras, 91% of the capture area was below 200 μm of error. The only locations that exceeded this threshold were at the extreme edges of the capture area, and no location had a mean error exceeding 1 mm. When measuring human kinematics with skin-mounted markers, uncertainty of marker placement relative to underlying skeletal features and soft tissue artifact produce errors that are orders of magnitude larger than the errors attributed to the camera system itself. Therefore, the accuracy of this OptiTrack optical motion capture system was found to be more than sufficient for measuring full-body human kinematics with skin-mounted markers in a large capture volume (>100 m³).     

A strategy to develop molecular markers applicable to a wide range of crosses for marker assisted selection in plant breeding: a case study on anthracnose disease resistance in lupin (Lupinus angustifolius L.)

A key challenge in marker-assisted selection (MAS) for molecular plant breeding is to develop markers linked to genes of interest which are applicable to multiple breeding populations. In this study representative F₂ plants from a cross Mandalup (resistant to anthracnose disease) × Quilinock (susceptible) of Lupinus angustifolius were used in DNA fingerprinting by Microsatellite-anchored Fragment Length Polymorphism (MFLP). Nine candidate MFLP markers linked to anthracnose resistance were identified, then ‘validated’ on 17 commercial cultivars. The number of “false positives” (showing resistant-allele band but lack of the R gene) for each of the nine candidate MFLP markers on the 17 cultivars ranged from 1 to 9. The candidate marker with least number of false positive was selected, sequenced, and was converted into a co-dominant, sequence-specific, simple PCR based marker suitable for routine implementation. Testing on 180 F₂ plants confirmed that the converted marker was linked to the R gene at 5.1 centiMorgan. The banding pattern of the converted marker was consistent with the disease phenotype on 23 out of the 24 cultivars. This marker, designated “AnManM1”, is now being used for MAS in the Australian lupin breeding program. We conclude that generation of multiple candidate markers, followed by a validation step to select the best marker before conversion to an implementable form is an efficient strategy to ensure wide applicability for MAS.     

大豆SSR标记辅助遗传背景选择的效果分析

本研究利用鲁豆4号回交转育的大豆种子脂氧酶缺失株系为材料,用IEF-PAGE鉴定大豆种子脂氧酶缺失基因,SSR标记进行遗传背景分析,通过遗传背景回复率相关分析,探索分子标记辅助遗传背景选择时所需的适宜的标记数和选择方式,期望获得可进一步回的鲁豆4号脂氧酶缺失株系。研究明确了大豆SSR标记辅助背景选择时适宜标记数目和选择方式,即先用少数标记初筛,选出遗传背景回复率较高的材料,再用适宜标记鉴定,随着世代递增,已恢复为轮回亲本的标记住点不再分析,逐代减少选择标记数目。获得了合有脂氧酶缺失基因,遗传背景与鲁豆4号差异较小的株系,可用鲁玉4号进一步回交,从而加速培育鲁豆4号脂氧酶缺失近等基因系。     

ParA resolvase catalyzes site-specific excision of DNA from the Arabidopsis genome

The small serine resolvase ParA from bacterial plasmids RK2 and RP4 catalyzes the recombination of two identical 133 bp recombination sites known as MRS. Previously, we reported that ParA is active in the fission yeast Schizosaccharomyces pombe. In this work, the parA recombinase gene was placed under the control of the Arabidopsis OXS3 promoter and introduced into Arabidopsis lines harboring a chromosomally integrated MRS-flanked target. The ParA recombinase excised the MRS-flanked DNA and the excision event was detected in subsequent generations in the absence of ParA, indicating germinal transmission of the excision event. The precise site-specific deletion by the ParA recombination system in planta demonstrates that the ParA recombinase can be used to remove transgenic DNA, such as selectable markers or other introduced transgenes that are no longer desired in the final product.     

Development of a set of highly polymorphic genomic microsatellites (gSSRs) in Sitka spruce (Picea sitchensis (Bong.) Carr.)

Robust, polymorphic microsatellite DNA markers (simple sequence repeats—SSRs) are valuable tools for a range of tree conservation and breeding applications. SSRs are routinely used in the study of population genetic structure and diversity, pedigree reconstruction and genetic linkage mapping. Their abundance in the genome, co-dominant inheritance and potential for cross-species amplification make microsatellites highly prized markers. This paper characterises 22 novel genomic polymorphic microsatellite loci for Sitka spruce (Picea sitchensis (Bong.) Carr.). Amplification of DNA from Sitka spruce material was carried out both with a set of unrelated trees to obtain diversity statistics for each locus, and with the progeny of a full-sib family to test simple Mendelian inheritance. Observed heterozygosity ranged from 0.38 to 0.91 and allele number per locus ranged from 6 to 21, with a mean of 12.2. In addition, the primer pairs were tested with DNA from Norway spruce (P. abies) and white spruce (P. glauca) to investigate their potential for cross-species amplification and ten loci amplified in all three species. The results from these genomic microsatellites are compared to data generated from microsatellites derived from Picea EST libraries. In summary, this novel, highly polymorphic markers represent a significant addition to the rapidly expanding Picea genomics tool-box. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

The multicompartmental p32/gClqR as a new target for antibody-based tumor targeting strategies

Tumor-associated cell surface antigens and tumor-associated vascular markers have been used as a target for cancer intervention strategies. However, both types of targets have limitations due to accessibility, low and/or heterogeneous expression, and presence of tumor-associated serum antigen. It has been previously reported that a mitochondrial/cell surface protein, p32/gC1qR, is the receptor for a tumor-homing peptide, LyP-1, which specifically recognizes an epitope in tumor cells, tumor lymphatics, and tumor-associated macrophages/myeloid cells. Using antibody phage technology, we have generated an anti-p32 human monoclonal antibody (2.15). The 2.15 antibody, expressed in single-chain fragment variable and in trimerbody format, was then characterized in vivo using mice grafted subcutaneously with MDA-MB-231 human breast cancers cells, revealing a highly selective tumor uptake. The intratumoral distribution of the antibody was consistent with the expression pattern of p32 in the surface of some clusters of cells. These results demonstrate the potential of p32 for antibody-based tumor targeting strategies and the utility of the 2.15 antibody as targeting moiety for the selective delivery of imaging and therapeutic agents to tumors.     

Regulation of sialyl Lewis antigen expression in colon cancer cells by sialidase NEU4

Sialyl Lewis antigens, sialyl Lewis a and sialyl Lewis x, are utilized as tumor markers, and their increase in cancer is associated with tumor progression by enhancement of cancer cell adhesion to endothelial E-selectin. However, regulation mechanisms are not fully understood. We previously demonstrated that NEU4 is the only sialidase efficiently acting on mucins and it is down-regulated in colon cancer. To elucidate the significance of NEU4 down-regulation, we investigated sialyl Lewis antigens as endogenous substrates for the sialidase. NEU4 was found to hydrolyze the antigens in vitro and decrease cell surface levels much more effectively than other sialidases. Western blot, thin layer chromatography, and metabolic inhibition studies of desialylation products revealed NEU4 to preferentially catalyze sialyl Lewis antigens expressed on O-glycans. Cell adhesion to and motility and growth on E-selectin were significantly reduced by NEU4. E-selectin stimulation of colon cancer cells enhanced cell motility through activation of the p38/Hsp27/actin reorganization pathway, whereas NEU4 attenuated the signaling. On immunocytochemical analysis, some NEU4 molecules were localized at cell surfaces. Under hypoxia conditions whereby the antigens were increased concomitantly with several sialyl- and fucosyltransferases, NEU4 expression was markedly decreased. These results suggest that NEU4 plays an important role in control of sialyl Lewis antigen expression and its impairment in colon cancer.