扇贝、对虾基因组BAC文库构建及其重要功能基因的初步筛选和分析

基因组大片段BAC文库是进行生物遗传学和基因组学研究必不可少的基础工具。为了深入开展栉孔扇贝（Chlamys farreri）和凡纳滨对虾（Litopenaeus vannamei）基因组学研究、阐明其基因组的结构与功能、图位克隆重要功能基因、构建高密度物理图谱并最终实现与已有遗传连锁图的整合，本研究在植物基因组BAC文库构建技术的基础上，针对海洋生物的特点进行了大胆的改革与尝试，最终成功构建了C. ferrari和L. vannamei两种重要海水养殖动物的基因组BAC文库。 本文构建的栉孔扇贝基因组BAC文库，由BamHI文库和MboI文库构成。其中BamHI文库含有73,728个BAC克隆，空载率约为1%，平均插入片段约为110 kb，覆盖栉孔扇贝单倍体基因组约8倍；MboI文库共有7,680个克隆组成，平均插入片段大小约为145 kb，插入率为100%，覆盖栉孔扇贝单倍体基因组约1.1倍。两个栉孔扇贝基因组BAC文库共由81,408个克隆组成，平均插入片段约为113 kb，覆盖率约为栉孔扇贝单倍体基因组大小的9.1倍。 将栉孔扇贝基因组BAC文库的192个384微孔培养板中的73,728个BAC克隆以4 x 4点阵形式制备了高密度DNA薄膜，用于对感兴趣的基因及DNA序列的筛选。高密度DNA薄膜的覆盖率约为栉孔扇贝单倍体基因组的8.3倍。针对栉孔扇贝先天免疫系统通路的6个重要功能基因，根据栉孔扇贝cDNA序列以及异缘物种DNA序列设计了Overgo探针。利用Overgo探针对高密度DNA薄膜杂交筛选的结果显示，平均每个基因检测到7.3个潜在阳性克隆。 本研究所构建的凡纳滨对虾基因组HindIII酶切BAC文库共有102,528个BAC克隆，存放于267个384微孔培养板中，平均插入片段大小约为101 kb，空载率约为5%，覆盖L. vannamei单倍体基因组约5倍。将其中240个384微孔培养板中的92,160个BAC克隆以4 x 4的矩阵排列形式制作了5张高密度凡纳滨对虾DNA薄膜，约覆盖整个对虾单倍体基因组的4.5倍。针对6个与对虾免疫、生殖生理有关的重要功能基因设计了Overgo探针，杂交筛选出20个阳性克隆，平均每个基因有3.3个潜在阳性克隆。 以上筛选结果不仅为进一步研究这些功能基因的结构与功能、表达与调控，揭示它们在扇贝和对虾以及其他近缘种的免疫系统、抗逆和生殖生理过程中的作用机理打下了基础，同时也间接验证了栉孔扇贝和凡纳滨对虾基因组BAC文库将成为基因筛选、基因的结构与功能分析、基因图位克隆、物理图谱构建以及大规模全基因组测序等方面的有力工具。

Construction of a Festuca pratensis BAC library for map-based cloning in IFestulolium substitution lines

Iain S. Donnison, Donal M. O Sullivan, Ann Thomas, Peter Canter, Beverley Moore, Ian Armstead, Howard Thomas, Keith J. Edwards and Ian P. King (2005). Construction of a Festuca pratensis BAC library for map-based cloning in Festulolium substitution lines. Theoretical and Applied Genetics, 110 (5) pp.846-851 Sponsorship: BBSRC;BBSRC RAE2008

A new look towards BAC-based array CGH through a comprehensive comparison with oligo-based array CGH

BACKGROUND: Currently, two main technologies are used for screening of DNA copy number; the BAC (Bacterial Artificial Chromosome) and the recently developed oligonucleotide-based CGH (Chromosomal Comparative Genomic Hybridization) arrays which are capable of detecting small genomic regions with amplification or deletion. The correlation as well as the discriminative power of these platforms has never been compared statistically on a significant set of human patient samples.

RESULTS: In this paper, we present an exhaustive comparison between the two CGH platforms, undertaken at two independent sites using the same batch of DNA from 19 advanced prostate cancers. The comparison was performed directly on the raw data and a significant correlation was found between the two platforms. The correlation was greatly improved when the data were averaged over large chromosomic regions using a segmentation algorithm. In addition, this analysis has enabled the development of a statistical model to discriminate BAC outliers that might indicate microevents. These microevents were validated by the oligo platform results.

CONCLUSION: This article presents a genome-wide statistical validation of the oligo array platform on a large set of patient samples and demonstrates statistically its superiority over the BAC platform for the Identification of chromosomic events. Taking advantage of a large set of human samples treated by the two technologies, a statistical model has been developed to show that the BAC platform could also detect microevents.

Comparaison des modèles HAR-RV et HAR-CJ sur les rendements de AMZN et BAC en 2010.

Rapport de recherche présenté à la Faculté des arts et des sciences en vue de l'obtention du grade de Maîtrise en sciences économiques.

L’exploration des incitatifs et des obstacles à entreprendre des études universitaires dans le cadre d’un cursus intégré DEC-BAC en sciences infirmières

Au Québec, les infirmières n’ont pas l’obligation de détenir un diplôme universitaire pour exercer leur profession. Celles qui choisissent le DEC en soins infirmiers peuvent obtenir le diplôme de baccalauréat en deux ans au lieu de trois en empruntant le cursus DEC-BAC en sciences infirmières. Or, ce n’est pas la majorité d’entre elles qui se prévalent de cette possibilité, alors que des études ont démontré les avantages d’avoir un plus grand nombre d’infirmières détenant un baccalauréat dans les milieux de soins (Aiken, Clarke, Cheung, Sloane & Silber, 2003; Kane, Shamliyan, Mueller, Duval & Wilt, 2007; Tourangeau, Cranley & Jeffs, 2006 ; Aiken et al., 2014). Le but de la présente étude était d’explorer les incitatifs et les obstacles à entreprendre des études universitaires dans le cadre du cursus intégré DEC-BAC en sciences infirmières. Cinquante-six nouvelles diplômées ayant complété un DEC en soins infirmiers au Québec ont répondu au questionnaire auto-administré électronique à questions ouvertes. Le cadre de référence guidant la recherche exploratoire était la théorie intermédiaire de la transition de Meleis, Sawyer, Im, Hilfinger Messias et Schumacher (2010). Suivant cette théorie, les données ont d’abord été regroupées par thèmes, soit « incitatifs » et « obstacles » puis par dimensions, à savoir si les thèmes relèvent de conditions au plan personnel, communautaire ou sociétal pour entreprendre des études universitaires. Il ressort de l’étude que les incitatifs majeurs à entreprendre des études universitaires se situent au plan communautaire : meilleures conditions de travail anticipées et développement professionnel. Les obstacles majeurs à entreprendre des études universitaires se situent au plan personnel, en particulier en lien avec une situation financière ou familiale défavorable. Nous avons aussi trouvé que le contexte québécois exerce une influence sur la décision d’entreprendre des études universitaires. Des recommandations ont été formulées pour les milieux de pratique et de formation, ainsi que pour la recherche.

Aportaciones al conocimiento de la geología de Bac Grillera (prepirineo, Girona)

The Bac Grillera nappe (Prepirenean of Girona) shows a stratigraphic section composed of two unconformi ty-bounded units. The lower unit is composed by rethian (marly limestones) and liassic (brechoid marly limestones and limestones). The upper unit (sandstones and marly limestones) belongs to the upper Cretaceous (Campanian)

Construction of a Festuca pratensis BAC library for map-based cloning in Festulolium substitution lines

Introgression in Festulolium is a potentially powerful tool to isolate genes for a large number of traits which differ between Festuca pratensis Huds. and Lolium perenne L. Not only are hybrids between the two species fertile, but the two genomes can be distinguished by genomic in situ hybridisation and a high frequency of recombination occurs between homoeologous chromosomes and chromosome segments. By a programme of introgression and a series of backcrosses, L. perenne lines have been produced which contain small F. pratensis substitutions. This material is a rich source of polymorphic markers targeted towards any trait carried on the F. pratensis substitution not observed in the L. perenne background. We describe here the construction of an F. pratensis BAC library, which establishes the basis of a map-based cloning strategy in L. perenne. The library contains 49,152 clones, with an average insert size of 112 kbp, providing coverage of 2.5 haploid genome equivalents. We have screened the library for eight amplified fragment length polymorphism (AFLP) derived markers known to be linked to an F. pratensis gene introgressed into L. perenne and conferring a staygreen phenotype as a consequence of a mutation in primary chlorophyll catabolism. While for four of the markers it was possible to identify bacterial artificial chromosome (BAC) clones, the other four AFLPs were too repetitive to enable reliable identification of locus-specific BACs. Moreover, when the four BACs were partially sequenced, no obvious coding regions could be identified. This contrasted to BACs identified using cDNA sequences, when multiple genes were identified on the same BAC.

A maize bacterial artificial chromosome (BAC) library from the European flint inbred line F2

We report here the construction and characterisation of a BAC library from the maize flint inbred line F2, widely used in European maize breeding programs. The library contains 86,858 clones with an average insert size of approximately 90 kb, giving approximately 3.2-times genome coverage. High-efficiency BAC cloning was achieved through the use of a single size selection for the high-molecular-weight genomic DNA, and co-transformation of the ligation with yeast tRNA to optimise transformation efficiency. Characterisation of the library showed that less than 0.5% of the clones contained no inserts, while 5.52% of clones consisted of chloroplast DNA. The library was gridded onto 29 nylon filters in a double-spotted 8 × 8 array, and screened by hybridisation with a number of single-copy and gene-family probes. A 3-dimensional DNA pooling scheme was used to allow rapid PCR screening of the library based on primer pairs from simple sequence repeat (SSR) and expressed sequence tag (EST) markers. Positive clones were obtained in all hybridisation and PCR screens carried out so far. Six BAC clones, which hybridised to a portion of the cloned Rp1-D rust resistance gene, were further characterised and found to form contigs covering most of this complex resistance locus.

Otimização e aperfeiçoamento BAC pra estudos envolvendo o trato gastrintestinal

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Modelo de estudo da motilidade gastrointestinal utilizando a eletromiografia (EMG) e a biosusceptometria AC (BAC) em ratos normais, desnutridos e diabéticos induzidas pela aloxana

Pós-graduação em Bases Gerais da Cirurgia - FMB

Construção de uma biblioteca BAC e avaliação de marcadores para caracterização de regiões alvo do genoma do búfalo

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Fontes hidrominerais do Brasil: componentes biologicamente ativos (BAC) naturais

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Consolidation of the genetic and cytogenetic maps of turbot (Scophthalmus maximus) using FISH with BAC clones

Bacterial artificial chromosomes (BAC) have been widely used for fluorescence in situ hybridization (FISH) mapping of chromosome landmarks in different organisms, including a few in teleosts. In this study, we used BAC-FISH to consolidate the previous genetic and cytogenetic maps of the turbot (Scophthalmus maximus), a commercially important pleuronectiform. The maps consisted of 24 linkage groups (LGs) but only 22 chromosomes. All turbot LGs were assigned to specific chromosomes using BAC probes obtained from a turbot 5x genomic BAC library. It consisted of 46,080 clones with inserts of at least 100 kb and < 5 % empty vectors. These BAC probes contained gene-derived or anonymous markers, most of them linked to quantitative trait loci (QTL) related to productive traits. BAC clones were mapped by FISH to unique marker-specific chromosomal positions, which showed a notable concordance with previous genetic mapping data. The two metacentric pairs were cytogenetically assigned to LG2 and LG16, and the nucleolar organizer region (NOR)-bearing pair was assigned to LG15. Double-color FISH assays enabled the consolidation of the turbot genetic map into 22 linkage groups by merging LG8 with LG18 and LG21 with LG24. In this work, a first-generation probe panel of BAC clones anchored to the turbot linkage and cytogenetical map was developed. It is a useful tool for chromosome traceability in turbot, but also relevant in the context of pleuronectiform karyotypes, which often show small hardly identifiable chromosomes. This panel will also be valuable for further integrative genomics of turbot within Pleuronectiformes and teleosts, especially for fine QTL mapping for aquaculture traits, comparative genomics, and whole-genome assembly.