Segmental Trans-Splicing Case Study

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Q1. Explain segmental trans-splicing (STS) and mention a long gene 8 kb or more, this strategy could be used for.
A1. Segmental trans-splicing concept is similar to that of trans-splicing, where exons of two different gene fragments are joined together. However, in STS it can be used to target gene sequences that are too large for a viral vector genetic capacity; for example the von Willbrand factor gene (8.6 kb) or the muscular dystrophy gene (11.0 kb). In STS a 5’exon of the gene is delivered in a vector and the 3’exon in a second vector where they are then joined together by the aid of a hybridization domain by spliceosomes to form the intact mRNA (O'Connor and Crystal, 2006).
Q2. Mention an example of tumor formed during stem cells pre-clinical
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In 2001, neural fetal stem cells were injected the brain and its fluid to a boy with ataxia telangiectasia (AT); a neurodegenerative disease where the region of the brain responsible for movement and speech degenerates. MRI scans carried out in 2005 revealed abnormal growths in brain and spinal cord. Masses removed surgically in 2006 where found to be tumor cells whose DNA did not match the DNA of the patient or his parents, but that of the donor fetus, concluding that it was caused by the injected stem cells (Amariglio et al., 2009).
Cancer stem cells are immortal cells found in tumors that have characteristics of a normal stem cells like the ability to differentiate into many cell types including that of the tumor, thus fueling its growth (Batlle and Clevers,
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Bacterial plasmid is circular DNA found in bacteria separate of its complete DNA in the nucleoide. It contains a multiple cloning site where the gene of interest is inserted in order to be expressed in transformed cells, an origin of replication and a selectable marker usually an antibiotic resistance gene which is used to select successfully transformed cells (Chin, n.d.). Mini-circle DNA is a smaller circular DNA excised from bacterial plasmid, contains promoter and reporter genes but does not contain any bacterial DNA sequences. It cannot replicate itself but its expression can last for 14 days (System Biosciences, n.d.). Mini-intronic plasmid DNA does not contain redundant bacterial DNA sequences as in mini-circle DNA therefore, both have an aided advantage of avoiding transgenic silencing. It also contains pUC origin of replication allowing the highest copy number (Morgan, 2014) and an RNA-OUT antisense selection rather than antibiotic selectable marker, making them safe and more efficient for gene therapy (Diecke et al., 2015), (Luke et al.,

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