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Thorsten Pfirrmann Ashwin Lokapally Claes Andréasson Per Ljungdahl Thomas Hollemann 《PloS one》2013,8(6)
Modern biology research requires simple techniques for efficient and restriction site-independent modification of genetic material. Classical cloning and mutagenesis strategies are limited by their dependency on restriction sites and the use of complementary primer pairs. Here, we describe the Single Oligonucleotide Mutagenesis and Cloning Approach (SOMA) that is independent of restriction sites and only requires a single mutagenic oligonucleotide to modify a plasmid. We demonstrate the broad application spectrum of SOMA with three examples. First, we present a novel plasmid that in a standardized and rapid fashion can be used as a template for SOMA to generate GFP-reporters. We successfully use such a reporter to assess the in vivo knock-down quality of morpholinos in Xenopus laevis embryos. In a second example, we show how to use a SOMA-based protocol for restriction-site independent cloning to generate chimeric proteins by domain swapping between the two human hRMD5a and hRMD5b isoforms. Last, we show that SOMA simplifies the generation of randomized single-site mutagenized gene libraries. As an example we random-mutagenize a single codon affecting the catalytic activity of the yeast Ssy5 endoprotease and identify a spectrum of tolerated and non-tolerated substitutions. Thus, SOMA represents a highly efficient alternative to classical cloning and mutagenesis strategies. 相似文献
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Milberg Per Bergman Karl-Olof Glimskär Anders Nilsson Sigrid Tälle Malin 《Plant Ecology》2020,221(7):577-594
Plant Ecology - Management of semi-natural grasslands is essential to retain the characteristic diversity of flora and fauna found in these habitats. To maintain, restore or recreate favourable... 相似文献
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Matheus E. Bianconi Luke T. Dunning Emma V. Curran Oriane Hidalgo Robyn F. Powell Sahr Mian Ilia J. Leitch Marjorie R. Lundgren Sophie Manzi Maria S. Vorontsova Guillaume Besnard Colin P. Osborne Jill K. Olofsson Pascal-Antoine Christin 《Proceedings. Biological sciences / The Royal Society》2020,287(1938)
C4 photosynthesis evolved multiple times independently in angiosperms, but most origins are relatively old so that the early events linked to photosynthetic diversification are blurred. The grass Alloteropsis semialata is an exception, as this species encompasses C4 and non-C4 populations. Using phylogenomics and population genomics, we infer the history of dispersal and secondary gene flow before, during and after photosynthetic divergence in A. semialata. We further analyse the genome composition of individuals with varied ploidy levels to establish the origins of polyploids in this species. Detailed organelle phylogenies indicate limited seed dispersal within the mountainous region of origin and the emergence of a C4 lineage after dispersal to warmer areas of lower elevation. Nuclear genome analyses highlight repeated secondary gene flow. In particular, the nuclear genome associated with the C4 phenotype was swept into a distantly related maternal lineage probably via unidirectional pollen flow. Multiple intraspecific allopolyploidy events mediated additional secondary genetic exchanges between photosynthetic types. Overall, our results show that limited dispersal and isolation allowed lineage divergence, with photosynthetic innovation happening after migration to new environments, and pollen-mediated gene flow led to the rapid spread of the derived C4 physiology away from its region of origin. 相似文献
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Özaslan Ahmet Kayhan Gülsüm İşeri Elvan Ergün Mehmet Ali Güney Esra Perçin Ferda Emriye 《Molecular biology reports》2021,48(11):7371-7378
Molecular Biology Reports - Copy number variants (CNVs) play a key role in the etiology of autism spectrum disorder (ASD). Therefore, recent guidelines recommend chromosomal microarrays (CMAs) as... 相似文献
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