Correct Answer : Option (D) - 1, 2 and 3
Note :
Gene editing of male and female germ cells
An alternative to the zygote/embryo approach is to perform gene modifications during early gametogenesis. In this manner, growing immature oocytes or sperm or even pre- cursor cells (primordial germ cells) can be gene targeted by using the CRISPR/Cas system, producing genetically corrected mature sperm or oocytes that subsequently can be used for ART. In the male germ cell line, spermatogonial stem cells (SSC) can be harvested more and more efficiently, and in vitro culture systems are being developed, also in the human, and optimized for efficient production of sperm in vitro. So far, animal models have indicated that SSC can be propagated as clones in culture and then transplanted back into the testis to generate mature and functional sperm . So a potential strategy would be to select SSC clones that have undergone correct genomic editing and are free from off-target mutations. These can then be transplanted to undergo final maturation in vivo. Alternatively, the gene edited SSC can be directly differentiated in vitro to mature gene-corrected sperm, to be used for IVF. Still, optimisation of in vitro culture systems is warranted, especially in the human, so that gene editing technologies can be employed safely and with high efficiency. In the female germ line, the oocyte is more easily accessible for genetic manipulation, but currently technical hurdles remain, such as the small number of oocytes that are available . It has been suggested that oogonia-like stem cells could be harvested, cultured and expanded followed by culture in vitro to the mature metaphase II stage . However, there is still controversy over the existence of such oocyte precursor cells in the female and the efficiency at which mature, developmentally competent oocytes can be derived from them.
Gene editing in zygotes or pre-implantation embryos
For germ-line modifications in experimental settings the genomic editing system is mostly injected into the cytoplasm or pronuclei of zygotes or into pre-implantation embryos, after which genetic screening is used to select the embryos with a corrected genomic pattern in the absence of detectable off-target genetic modifications. Should this turn out to be safe and effective, then similar applications in the clinic are conceivable. Subsequently, prenatal testing using either cell-free foetal DNA from the pregnant woman’s blood or one of the more invasive methods (chorionic villus sampling or amniocentesis), could in theory verify whether or not a foetus shows molecular or genomic mosaicism. Mosaic embryos arise as a result of inefficient cutting of the nucleases or inaccurate DNA repair before the embryo has reached the stage of cleavage. The pre-implantation embryo stage is generally not favoured for genome editing as it would most likely lead to a mosaic individual and possibly to more unforeseen detrimental effects. Various studies in different animal models have demonstrated the feasibility of gene editing in animals at the zygote stage.Hence, statements 1 and 2 are correct.
The controversial idea of growing human organs in host animals has passed through a reality check. Despite recent successes at growing mouse organs in rats, using the same trick to grow human organs in larger animals such as pigs is a long way off, new research shows. The researchers tried combining human induced pluripotent stem cells (reprogrammed adult cells that have regained the characteristics of embryonic cells) with pig embryos. After they implanted the chimeras into surrogate mothers, they let the embryos develop for just 3 or 4 weeks, to check whether and where the human cells were contributing. The researchers implanted more than 2000 human-pig chimeric embryos into 41 surrogate sows, resulting in 18 pregnancies and 186 embryos a month later. However, many of the embryos were much smaller than normal and seemed to grow more slowly, the group reports today in Cell. Hence, so far there has not been success in growing human organs in pigs. But this experiment shows that human induced pluripotent cells can be injected into pigs embryo. Hence statement 3 is also correct.