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...separately introduce each mutation independently into different mice, and then these strains should be crossed...of the mutations engineered into each of their parents. Here’s how to cross mice, using a simple example...Mendalian patterns of inheritance: 1. Genotype your parental mice In order to ensure that your genetic modification...generation, your breeding scheme should start with parental mice that are each homozygous for one of the two...targeting vector or gene of interest. 2. Breed your parental mice to generate F1 Once you’ve performed your...desired combination of mutations. Assuming your parental mice were all homozygous for your genes of interest...breeding heterozygous mice. In this case, your parental mouse should be hemizygous at the appropriate ...

... a virus enters the cell. Different serotypes utilize a number of different receptors, co-receptors, and...burdensome in the brain where many different cell types, or even different phenotypes of the same cell type...working in several different systems or want to transduce a large number of different cells. Promoters ...doi.org/10.3389/fnmol.2014.00076 References and Resources References Aschauer, D. F., Kreuz, S., & Rumpel...validation and optimization. Although there are different ways to ultimately achieve success, we find it...variables.    Serotypes Serotype refers to the different protein compositions of AAV capsids, which determine... system or cell type can completely fail in a different system (Issa et al., 2023). This can be particularly...

...antibodies are all of different species/isotypes and that each secondary has a different color fluorophore...a comment below. Happy imaging! References and Resources References Andrews, N. P., Boeckman, J. X., ...mixture of secondary antibodies, and image all the different colors in one session – fast and easy! Just make...Trimmer Lab, using recombinant antibodies with different isotypes. (A) Neocortex labeled with anti-pan-...make sure you are using primary antibodies from different species or isotypes, so that the secondary antibodies...to align the images of each labeled target as different color channels and analyze the results. Depending...you might need to quantify the intensity of the different signals in a certain region or calculate a colocalization...

...resolving gel. A stacking gel typically has a different ionic strength and lower pH and acrylamide content than...validated for immunoblotting. Different immunoassays detect proteins in different states. For example, in immunoprecipitation... other antibody topics. Resources and references References Aydin S (2015) A short history, principles...to detect the presence or absence of a protein (Renart et al., 1979, Towbin et al., 1979, Burnett et al...the size of the protein of interest. An electric current applied to the gel causes the negatively charged...repeatedly stripped of an antibody and reprobed with different antibodies allowing multiple targets to be assessed...” together with blotting paper and an electric current is applied causing proteins to migrate out of the...

...common AAV serotypes used for different tissues. Created with BioRender.com. Beyond the identified serotypes...Your AAVancement could be next!   References and Resources References Braun, C. J., Adames, A. C., Saur...allows for a wide variety of screens, including interference, activation, and knockout. Most CRISPR screens...CRISPR screens can be carried out using a few different methods — in vivo, in vitro, and ex vivo. The ...of in vitro AAV CRISPR screens. Created with BioRender.com. The in vitro option requires a bit less...mouse tissues. Safety AAV and lentivirus have different safety profiles that often affect which is chosen...in a variety of capsids. To date, at least 12 different capsids, or serotypes, have been identified in...

...how to spend my grant money (Can I attend this conference?) and with my lab mates on which lab jobs needed... also negotiate with friends, partners, and/or children about everything from what you’re going to eat...another author can present the work at the next big conference - Is one of those options more important to you...are their expectations/needs and how are they different from mine? What equipment purchases do we prioritize...she’s trying to get some data for an upcoming conference deadline. Instead of arguing over who gets the... takes the earlier slot agrees to pick up both children while the other person uses an available after-hours... or coursework. First, you have to develop an awareness of when you are negotiating. Then take a moment...

...software, you then generate a trendline for the data. The appropriate trendline varies between assays and ...linear trendlines while those for ELISAs and other enzymatic assays are sigmoidal. Once the trendline is ...non-linear and you should use a different method of analysis. Different fields and assays have varying ...well by vortexing or inversion. Created with BioRender.com.   When choosing a standard, make sure that...plotted against its OD560 measurements and the trendline y = 0.6561x + 0.019 calculated. The concentration... an unknown sample is determined by using the trendline to extrapolate its x value, 0.963 mg/mL, from ...and measures how well the data points fit the trendline. R2 ranges from 0 to 1 with 1 being a perfect ...

... if you want to be an entrepreneur in the health sciences?  Is this a different story altogether? The ...Being an entrepreneur is difficult; it requires an immense amount of self-knowledge and an unwavering...become something tangible.  When you think of entrepreneurship in the 21st century, who do you think of?  ...with innovative ideas in all facets of the entrepreneurial process in order to develop those ideas into... about 3 project proposals per month.  We are currently focused on two projects, including one led by ...lessons I learned while becoming a health science entrepreneur at 22. Start with your network You've spent ...viability question. The miLab you see today is far different than what was originally planned, due in large...

...when interpreting the outcome.    References and resources References Aschauer DF, Kreuz S, Rumpel S (2013...2, and 5 Display Differential Efficiency and Cell Tropism after Delivery to Different Regions of the Central... recognized by Cre, but they are comprised of different sequences (loxp and lox2272) and can not recombine...transgene from reverting. Figure created with BioRender.com   Causes of recombination and leaky expression... flank a transgene (homologous pairs) or used different scrambled sequences to flank the transgenes (non-homologous... off-target expression. Figure created with BioRender.com   Strategies for optimizing expression So...  The AAV serotype used can exhibit profound differences in tropism between cell types (astrocytes, microglia...

...from that sequence preferences must largely be ignored. Created with BioRender.com. The hammer...exact location of the TSS. Different databases annotate the TSS in different ways, and it has been shown...phenotype of interest. This blog post will discuss differences between these approaches, and provide updates...Doench et al., 2017, and Hanna et al., 2020) and references therein.   {% module_block module "widget_fcf9eab0...experiment with CRISPR The hammer, the jigsaw, and the wrench are all great tools, but which one you use, of ...importance (more on this below). Created with BioRender.com. The jigsaw: Editing by HDR, base editing...few potential gRNAs. While SpCas9, with a PAM preference of NGG, is still the most widely-used Cas enzyme...

... cell. Differential expression of transcription factors, which recognize and bind to different DNA motifs...the genome based on their differential chromatin accessibility across different cell types. Most enhancers...department of Molecular Genetics. References and Resources References Ben-Simon, Y., Hooper, M., Narayan...usually hear this phrase when discussing raising children, it can also be applied to research. Sharing knowledge...resources to generate highly specific tools to target different cell populations and regions of the brain. This...transcriptional regulation by an enhancer. Created with BioRender.com.   Developing AAV enhancer tools for brain...only in target cells. This pattern is termed “differential DNA accessibility,” and AIBS uses it to find...

...Cary Valley, and Holly McQueary References and Resources References Niedringhaus, T. P., Milanova, D.... Sanger sequencing technology. Created with BioRender.com. Sanger sequencing was considered the gold...short-read sequencing technology. Created with BioRender.com. After sequencing results are deconvoluted...identify the bases (and, if necessary, separate different samples), the resulting data, or reads, are essentially... the flow cell membrane and have an electrical current running through them. DNA or RNA is pulled through...nucleotide passes through the nanopore, the electrical current recorded by the instrument changes. Base-calling...Nanopore sequencing technology. Created with BioRender.com. Nanopore can provide longer read lengths...

...plasmids are compatible, different CBPs bind to each plasmid type, and different NTPases separate the plasmid...plasmids at Addgene Read our Molecular Biology Reference References  del Solar G, Giraldo R, Ruiz-Echevarría...incompatibility is defined as the inability of different plasmids to be maintained in one bacterial cell...al., 1998).  In cases of compatible plasmids, different negative regulation systems control plasmid replication...regulation system cannot distinguish between the different plasmids. The concentration of negative regulation...diffusion) to make this happen.  There are several different versions of the plasmid partitioning system in...identification issue: the CBPs in the cell can’t tell the difference between plasmids if they have the same centromere-like...

...for Neuroscience 2017 conference in Washington DC (#SfN17). This massive conference attracted over 30,000...developed in your current work to make new insights in neuroscience. Even if you're currently a synthetic biologist...-depth exposure to neuroscience prior to this conference was a rotation in Qiao Zhou’s lab in graduate...school where I worked on a project aiming to dedifferentiate mouse brain cells into neural stem cells. I...biologists. This was not at all the case. The conference featured people who work in sociology, behavior... Collider and which featured a talk about the different parts of the brain stimulated by stories. In retrospect...neuroscientists are using techniques from across different disciplines to better understand neurobiology ...

...through a tiny pore by measuring differences in ion flow caused by the different electrochemical properties ...Harvard University currently studying the serotonergic system and behavior.     References 1. Southern, Edwin...all cases, the result can be quantified by the strength of the signal in comparison to a control sample...results of cloning or knockout experiments to forensics and clinical diagnostics. One example of southern...standard PCR. This allows the researcher to compare different DNA sequences to see which is more abundant in...content by comparing the fragments created by different restriction enzyme digests. Incubating a DNA sample...incorporation of a ddNTP. Researchers perform four different reactions, one for each radiolabelled ddNTP, and...

...updated in 2023 by Susanna Stroik.  References and Resources References Yuxuan Wu et al. "Correction of a...of the CRISPR-based editing therapies that are currently approved or have made significant progress in ...cells and delivering them to the patient.   Current CRISPR Therapies Cancer and CAR-T therapy The FDA...therapy pipelines (viral), and researchers are currently working on giving this treatment a CRISPR spin...leukemia, lymphoma, and some solid tumors are currently in early-phase clinical trials and have shown ...quite the opposite aim: to destroy. One treatment currently in clinical trials includes Cas3 (the shredder... into many, many pieces. Locus Biosciences is currently running this trial, with more updates expected...

...several different markers, it can also be used to run multiplex assays, allowing the reference sequence...primer  Molecular Biology Reference Molecular Cloning Techniques References Bell AD, Usher CL, McCarroll...each cycle. The raw data is then normalized to a reference sequence present in the same amount in all samples...curve. For CNV assays, it is recommended to use a reference sample with two copies of the gene (Ma, 2014.)...copy numbers, though it has been shown to identify trends correctly (Muhamad, 2011;  Providenti, 2006). Its...probe set for both the gene of interest and the reference sequence. At the end of the run, each droplet ...positive droplets for the gene of interest and reference gene, copy number can be determined through concentration...

... of the plasmids in our repository with known reference sequences, such as full plasmid sequences provided...BLAST, which includes a simple explanation of the different BLAST programs, databases, and elements of the...The blastp and blastx programs are optimized differently and you may want to select one (or both) depending...you want to verify. We will delve into these differences below. Optimizing blastn searches On the Standard... a Sanger sequencing result to a single known reference sequence or to a BLAST sequence database. If you...two or more sequences” checkbox and paste your reference sequence into the Subject Sequence box that appears...BLAST searches. If you do not know the exact reference sequence for your result, choose one of the BLAST...

... in mice Many systemic capsids express differently in different mouse strains, a phenomenon that we are...And these differences occur not just between mice and NHPs; some capsids show both different efficacy ...from AAV9, the vectors have to be administered concurrently to minimize the impact of neutralizing antibodies...determined by the capsid and promoter/enhancer strengths in the cell type of interest as well as the type...from Challis et al. 2022. Image created with BioRender.com. Abbreviations: AAV, adeno-associated virus;...Organism Mouse Strain-Specific Tropism?  Reference AAV-PHP.B* CNS (broad) Mice, rats Yes...Mechanisms of crossing the BBB are indicated in parentheses in the"Mouse Strain-Specific Tropism?" column...

... that differences in gene expression could explain why some ancestry groups were differentially dependent...ancestry-associated genes.   What does the reference genome actually reference?  The CRISPR guides used in the screens...done in this preprint.   Resources and references References Germline variation contributes to false...racial bias, which still impacts and influences current research. Understanding the depth and breadth of... were designed to have a perfect match to the reference genome – even single mismatches within a CRISPR...introduce significant bias into the results. But the reference genome does not represent the diversity of genetic...understand how the use of a primarily European reference genome could affect their analysis. Their work...