by Aidan O’Brien, Denis C. Bauer, Gaetan Burgio Genome editing through the development of CRISPR (Clustered Regularly Interspaced Short Palindromic Repeat)–Cas technology has revolutionized many fields in biology. Beyond Cas9 nucleases, Cas12a (formerly Cpf1) has emerged as a promising alternative to Cas9 for editing AT-rich genomes. Despite the promises, guide RNA efficiency prediction through computational tools search still lacks accuracy. Through a computational meta-analysis, here we report that Cas12a target and off-target cleavage behavior are a factor of nucleotide bias combined with n ..read more

by Shubhada Shenai, Gita Nataraj, Minal Jinwal, Akhil S. Thekke Purakkal, Rajashree Sen, Nayana Ingole, Trupti Mathure, Sanjay Sarin, Sarabjit S. Chadha Background Tata MD CHECK SARS-CoV-2 kit 1.0, a CRISPR based reverse transcription PCR (TMC-CRISPR) test was approved by Indian Council of Medical Research (ICMR) for COVID-19 diagnosis in India. To determine the potential for rapid roll-out of this test, we conducted performance characteristic and an operational feasibility assessment (OFA) at a tertiary care setting. Intervention The study was conducted at an ICMR approved COVID-19 RT-PCR lab ..read more

by Omer Benjakob, Olha Guley, Jean-Marc Sevin, Leo Blondel, Ariane Augustoni, Matthieu Collet, Louise Jouveshomme, Roy Amit, Ariel Linder, Rona Aviram Rapid developments and methodological divides hinder the study of how scientific knowledge accumulates, consolidates and transfers to the public sphere. Our work proposes using Wikipedia, the online encyclopedia, as a historiographical source for contemporary science. We chose the high-profile field of gene editing as our test case, performing a historical analysis of the English-language Wikipedia articles on CRISPR. Using a mixed-method approa ..read more

by Daniel A. Kuppers, Jonathan Linton, Sergio Ortiz Espinosa, Kelly M. McKenna, Anthony Rongvaux, Patrick J. Paddison Human CD34+ hematopoietic stem and progenitor cells (HSPCs) are a standard source of cells for clinical HSC transplantations as well as experimental xenotransplantation to generate “humanized mice”. To further extend the range of applications of these humanized mice, we developed a protocol to efficiently edit the genomes of human CD34+ HSPCs before transplantation. In the past, manipulating HSPCs has been complicated by the fact that they are inherently difficult to transduce ..read more

by Mohammadreza Paraan, Mohamed Nasef, Lucy Chou-Zheng, Sarah A. Khweis, Allyn J. Schoeffler, Asma Hatoum-Aslan, Scott M. Stagg, Jack A. Dunkle Type III CRISPR-Cas systems employ multiprotein effector complexes bound to small CRISPR RNAs (crRNAs) to detect foreign RNA transcripts and elicit a complex immune response that leads to the destruction of invading RNA and DNA. Type III systems are among the most widespread in nature, and emerging interest in harnessing these systems for biotechnology applications highlights the need for detailed structural analyses of representatives from diverse org ..read more

by Ikuko Nakajima, Hiroyuki Kawahigashi, Chikako Nishitani, Akifumi Azuma, Takashi Haji, Seiichi Toki, Masaki Endo Transposition of transposable elements affect expression levels, splicing and epigenetic status, and function of genes located in, or near, the inserted/excised locus. For example, in grape, presence of the Gret1 retrotransposon in the promoter region of the VvMYBA1a allele at the VvMYBA1 locus suppress the expression of the VvMYBA1 transcription factor gene for the anthocyanin biosynthesis and this transposon insertion is responsible for the green berry skin color of Vitis labras ..read more

by Aya Bridgeland, Sudip Biswas, Nikolaos Tsakirpaloglou, Michael J. Thomson, Endang M. Septiningsih Cowpea (Vigna unguiculata) is a legume staple widely grown across Sub-Saharan Africa and other tropical and sub-tropical regions. Considering projected climate change and global population increases, cowpea’s adaptation to hot climates, resistance to drought, and nitrogen-fixing capabilities make it an especially attractive crop for facing future challenges. Despite these beneficial traits, efficient varietal improvement is challenging in cowpea due to its recalcitrance to transformation and lo ..read more

by Pouiré Yaméogo, Nathalie Majeau, Cedric Happi Mbakam, Jacques P. Tremblay The small size of CjCas9 can make easier its vectorization for in vivo gene therapy. However, compared to the SpCas9, the CjCas9 is, in general, less efficient to generate indels in target genes. The factors that affect its efficacity are not yet determined. We observed that the CjCas9 protein expressed in HEK293T cells after transfection of this transgene under a CMV promoter was much lower than the SpCas9 protein in the same conditions. We thus evaluated the effect of proteasome inhibitors on CjCas9 protein stabilit ..read more

by Emily K. Matozel, Stephen Parziale, Allen C. Price A protein roadblock forms when a protein binds DNA and hinders translocation of other DNA binding proteins. These roadblocks can have significant effects on gene expression and regulation as well as DNA binding. Experimental methods for studying the effects of such roadblocks often target endogenous sites or introduce non-variable specific sites into DNAs to create binding sites for artificially introduced protein roadblocks. In this work, we describe a method to create programmable roadblocks using dCas9, a cleavage deficient mutant of the ..read more

by Shymaa Enany, Samira Zakeer, Aya A. Diab, Usama Bakry, Ahmed A. Sayed Klebsiella pneumoniae is considered a threat to public health especially due to multidrug resistance emergence. It is largely oligoclonal based on multi-locus sequence typing (MLST); in Egypt, ST 627 was recently detected. Despites the global dissemination of this ST, there is still paucity of information about it. Herein, we used 4 K. pneumoniae ST627 for whole genome sequencing utilizing an Illumina MiSeq platform. Genome sequences were examined for resistance and virulence determinants, capsular types, plasmids, insert ..read more