Mouse Model And Sample Size

Every few years, advancements in technology alter the way scientists do their work. Recently CRISPR-Cas9, a RNA useful for working organisms in the animal kingdom has proven itself beneficial on a gene-editing platform. After performing many abortive attempts to manipulate gene function, including homologous recombination and RNA interference, scientists have finally had a breakthrough with CRISPR-Cas9.

CRISPR is versatile in that any target sequence can be modified by simply altering the gRNA sequence. In addition, multiple genes can be edited at the same time with great specificity (Cong 88-89). The convenience and accessibility of CRISPR resourced have also allowed thousands of laboratories worldwide to study CRISPR in different ways, which has broadened the horizons of its biomedical and clinical implications (Collins et al 259). Overall, the ease and simplicity of CRISPR technology has allowed for a rapid increase in the understanding of genome editing, which will allow CRISPR to revolutionize how certain conditions will be treated.

CRISPR-Cas9 is a unique technology that can be used to edit the human genome (1). Compared to other techniques of editing DNA, CRISPR and Cas9 are cheaper, faster, and more accurate. CRISPR-Cas9 gives geneticists as well as medical researchers the ability to edit specific parts of the genome by the removal, adding, or altering of certain sections of DNA sequences (1). CRISPR stands for Clustered Regularly Interspaced Short Palindromic Repeats and is a part of bacterial defense system (2). The two key molecules of CRISPR-Cas9 are Cas9 (an enzyme that can cut DNA at specific locations) and guide RNA (a small piece of RNA sequence that guides where the Cas9 should cut) (1). CRISPR is a crucial component

The usage of nucleases is rapidly growing in an effort to save millions of lives worldwide; recently, gene editing tool was used to treat little girl’s cancer. Targeted genome editing is the most efficient, and effective way to treat cancer. Generally speaking CRISPR/Cas9 a powerful gene-editing tool should be considered to help the millions people that are losing their lives daily. As a matter of fact, researchers have developed three different kinds of gene-editing tools known as ZFN, TALEN, and newest and very effective CRISPR/Cas9. In the final analysis, CRISPR/Cas9 is the gene-editing tool that must be used in treatment of millions of people who are living with cancer. Recently, experiments were performed using CRISPR/Cas9, for instance editing the mushroom, modified human embryos, targeted mutagenesis, and genome editing in sea urchin embryos indicate high levels of efficiency, accuracy, and is opening door to treating deadly

The Cas9 follows the guide RNA to the same location in the DNA sequence and makes a cut across both strands of the DNA. At this stage the cell recognises that the DNA is damaged and tries to repair it. Scientists can use the DNA repair machinery to introduce changes to one or more genes in the genome of a cell of interest. According to a team of scientists working on CRISPR, “The guide RNA is designed to find and bind to a specific sequence in the DNA. The guide RNA has RNA bases that are complementary to those of the target DNA sequence in the genome. This means that, at least in theory, the guide RNA will only bind to the target sequence and no other regions of the genome.” The CRISPR-Cas9 has a gene editing

The CRISPR-Cas9 complex is derived from the immune system of bacterial cells and also contains repurposed exogenous RNA’s responsible for editing the human genome.1 To edit a gene within the genome, researchers add a CRISPR RNA (crRNA), which is complementary to the DNA code of interest and is responsible for binding. The crRNA is engineered to be extremely specific to the code of interest. A trans-activating CRISPR RNA (tracrRNA) is also necessary for guiding all the pieces of the complex together to carry out the function of editing. The repurposed CRISPR-Cas9 complex contains two nucleases, RuvC and HNH, which perform noncomplementary strand cleavage and complementary

CRISPR Cas9 is a gene editing tool that can be used to edit, delete, and change parts of the genome. What makes CRISPR Cas9 different from other gene editing techniques such as Zinc Finger Nuclease (ZFN) or Transcription Activator-Like Effector Nucleases (TALEN) is its targeting efficiency ability. For example, ZFNs and TALENs could only reach targeting efficiencies from 1% to 50% in human cells. On the other hand, CRISPR Cas9 had a much higher rate at greater than 70% in Zebrafish and plants. (Reis, 2014). CRISPR Cas9 is still being heavily researched by scientists today, because although many advancements have been made since its discovery in 1993, it is far from ready to be used commonly. Successful studies and experiments with this gene

CRISPR-Cas9 has recently become a popular set of tools for genetic engineering. By targeting specific DNA sequences, biotechnology can edit portions of organism’s genome by adding, replacing, or deleting sequences of DNA. Biotechnology also allows genes to be transferred from one organisms to another. The system uses a modified bacterial protein and a RNA to guide it to a specific DNA sequence. Scientist have also found ways to use these tools to send proteins to DNA targets to remove or add genes. When there is a significant error during repair, an entire sequence can become altered and the CRISPR system can study what happened to the cell which this error occurred in. The system has opened up new possibilities for discovering all areas of

The CRISPR-Cas (clustered regularly interspaced short palindromic repeat-CRISPR associated protein) system is a defense mechanism found in bacteria and archaea to target and inactivate foreign DNA materials such as viral DNA. The foreign DNA is integrated into the CRISPR array. The inserted DNA, known as a spacer, is transcribed from CRISPR loci and translated into short CRISPR RNAs (crRNAs). The crRNA, also known as a guide RNA (gRNA), forms a complex with Cas9. This RNA-Cas9 complex scans for invading DNA materials complementary to the gRNA and cleaves it by Cas9, a protein encoded by CRISPR loci. Cas9 consists of an RNA binding domain, nuclease lobe for DNA cleavage, and protospacer adjacent motif (PAM) interacting sites recognizing the

Hsu, P., Lander, E., & Zhang, F. (2014). Development and Applications of CRISPR-Cas9 for Genome Engineering. Cell, 157(6), 1262-1278. http://dx.doi.org/10.1016/j.cell.2014.05.010

After the growth of a teratoma, the director of the laboratory, Dr. Clarence Cook Little, encouraged Stevens to continue in this line of research, including exploratory dissections (Kelly 23). Then he found that there were some cells that were normally found in the cardiac muscle and some other cells that did not belong there (Kelly 23). After, that discovery they named it strain "mouse 129". Stevens found that many other mice were developing teratomas (Kelly 23). This all indicated that all these mice generated a gene that would develop teratomas. Also, most of these cells had undifferentiated and differentiated cells (Kelly 23).

Estrogen is the primary female sex hormone and is responsible for the development and regulation of the female reproductive system and of secondary sex characteristics. Produced primarily by the ovaries, estrogen binds to and activates estrogen receptor beta (ER)- a nuclear receptor. Estrogen plays an important role in the hypothalamic-pituitary-gonadal (HPG) axis by acting on gonadotropin-releasing hormone (GnRH) neurons located within the hypothalamus. In females, a cyclic reversal of estrogen feedback produces a positive feedback loop at both the hypothalamic and pituitary levels. This is coupled with an increase in pituitary sensitivity to GnRH, which produces the massive surge of luteinizing hormone (LH) that triggers ovulation. However, LH is not only secreted during this surge- it is secreted in a pulsatile manner otherwise. This paper will focus on the implications on LH pulsatility by knocking out ER, as well as by removing the ovaries.

Thank to my undergraduate education in biology at the Complutense University of Madrid, I gained a solid background in cell and molecular biology, biomedicine and neurobiology. Afterwards, as a master student, I have increased my academic training in neurosciences at the VU University Amsterdam in a highly international and intellectually challenging environment. Due to my participation in numerous seminars, journal clubs and poster markets, I have developed a strong critical thinking and remarkable communication skills. I have complemented my education with a course in mice handling and safety

Figure 6 : A graph showing the expected body weight of mice successfully treated by CRISPR/cas gene editing and that expected if the the mice still exhibit SMA symptoms.

Clustered regularly interspaced short palindromic repeats (CRISPR) is a microbial adaptive immune system which utilizes an RNA guided Cas9 endonuclease to specifically target and destroy viral DNA via double stranded breaks (Cong et al., 2013; Hsu et al., 2014). Researchers discovered that they could manipulate the guide RNA (gRNA) to deliver Cas9 to a DNA sequence of interest within the human genome. This enables Cas9 to induce site specific double stranded breaks (Hsu et al., 2014). Donor DNA sequences containing fully functional genes ending in DNA that is homologous to the cleaved ends can then be incorporated into the break via homologous directed repair (HDR) by the cell’s own DNA-repair mechanism (Mali et al., 2013). In this way,