RESUMO
Agricultural biotechnology is limited by the inefficiencies of conventional random mutagenesis and transgenesis. Because targeted genome modification in plants has been intractable, plant trait engineering remains a laborious, time-consuming and unpredictable undertaking. Here we report a broadly applicable, versatile solution to this problem: the use of designed zinc-finger nucleases (ZFNs) that induce a double-stranded break at their target locus. We describe the use of ZFNs to modify endogenous loci in plants of the crop species Zea mays. We show that simultaneous expression of ZFNs and delivery of a simple heterologous donor molecule leads to precise targeted addition of an herbicide-tolerance gene at the intended locus in a significant number of isolated events. ZFN-modified maize plants faithfully transmit these genetic changes to the next generation. Insertional disruption of one target locus, IPK1, results in both herbicide tolerance and the expected alteration of the inositol phosphate profile in developing seeds. ZFNs can be used in any plant species amenable to DNA delivery; our results therefore establish a new strategy for plant genetic manipulation in basic science and agricultural applications.
Assuntos
Biotecnologia/métodos , Desoxirribonucleases/química , Desoxirribonucleases/metabolismo , Marcação de Genes/métodos , Genoma de Planta/genética , Zea mays/genética , Dedos de Zinco , Desoxirribonucleases/genética , Alimentos Geneticamente Modificados , Genes de Plantas/genética , Resistência a Herbicidas/genética , Herbicidas/farmacologia , Hereditariedade , Fosfatos de Inositol/metabolismo , Mutagênese Sítio-Dirigida/métodos , Plantas Geneticamente Modificadas , Recombinação Genética/genética , Reprodutibilidade dos TestesRESUMO
The course-based research experience (CRE) with its documented educational benefits is increasingly being implemented in science, technology, engineering, and mathematics education. This article reports on a study that was done over a period of 3 years to explicate the instructional processes involved in teaching an undergraduate CRE. One hundred and two instructors from the established and large multi-institutional SEA-PHAGES program were surveyed for their understanding of the aims and practices of CRE teaching. This was followed by large-scale feedback sessions with the cohort of instructors at the annual SEA Faculty Meeting and subsequently with a small focus group of expert CRE instructors. Using a qualitative content analysis approach, the survey data were analyzed for the aims of inquiry instruction and pedagogical practices used to achieve these goals. The results characterize CRE inquiry teaching as involving three instructional models: 1) being a scientist and generating data; 2) teaching procedural knowledge; and 3) fostering project ownership. Each of these models is explicated and visualized in terms of the specific pedagogical practices and their relationships. The models present a complex picture of the ways in which CRE instruction is conducted on a daily basis and can inform instructors and institutions new to CRE teaching.
Assuntos
Modelos Educacionais , Estudantes , Engenharia , Docentes , Humanos , Matemática , EnsinoRESUMO
Strains of Drosophila melanogaster with resistance to the insecticides spinosyn A, spinosad, and spinetoram were produced by chemical mutagenesis. These spinosyn-resistant strains were not cross-resistant to other insecticides. The two strains that were initially characterized were subsequently found to have mutations in the gene encoding the nicotinic acetylcholine receptor (nAChR) subunit Dalpha6. Subsequently, additional spinosyn-resistant alleles were generated by chemical mutagenesis and were also found to have mutations in the gene encoding Dalpha6, providing convincing evidence that Dalpha6 is a target site for the spinosyns in D. melanogaster. Although a spinosyn-sensitive receptor could not be generated in Xenopus laevis oocytes simply by expressing Dalpha6 alone, co-expression of Dalpha6 with an additional nAChR subunit, Dalpha5, and the chaperone protein ric-3 resulted in an acetylcholine- and spinosyn-sensitive receptor with the pharmacological properties anticipated for a native nAChR.
Assuntos
Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Resistência a Medicamentos/genética , Inseticidas/farmacologia , Macrolídeos/farmacologia , Receptores Nicotínicos/genética , Receptores Nicotínicos/metabolismo , Animais , Chaperoninas/genética , Chaperoninas/metabolismo , Drosophila melanogaster , Combinação de Medicamentos , Resistência a Medicamentos/efeitos dos fármacos , Expressão Gênica , Mutação , Oócitos/citologia , Oócitos/metabolismo , Xenopus laevisRESUMO
SUMMARY Quinoxyfen is a protectant fungicide which controls powdery mildew diseases by interfering with germination and/or appressorium formation. Mutants of barley powdery mildew, Blumeria graminis f.sp. hordei, which are resistant to quinoxyfen produce fewer conidia, which germinate and form appressoria more promiscuously than do the prolific numbers of wild-type spores. This suggests that resistance bypasses host recognition signals. RT-PCR profiles of signal transduction genes, recorded during wild-type germling morphogenesis, reveals that quinoxyfen alters the accumulation of Protein Kinase C (pkc), pkc-like and catalytic subunit of Protein Kinase A (cpka) transcripts. Differential display-reverse transcription PCR identified a gene transcript in wild-type conidia that was absent, or much less abundant, in conidia from quinoxyfen-resistant mutants. This mRNA was not detectable 24 h after wild-type conidia were inoculated on to barley. It encodes a GTPase activating protein (GAP), which may interact with a small molecular weight Ras-type GTP binding protein. In the presence of quinoxyfen, the gap mRNA remains throughout germling morphogenesis. The involvement of GAP in resistance suggests that quinoxyfen inhibits mildew infection by disrupting early cell signalling events.