Biopharming has been gaining much more attention in the last few decades. Biopharming is the production and use of transgenic animals and plants to produce pharmaceutical substances for use in animals and humans ^[1]. Advancements in biopharming have laid a foundation to produce phenotypes such as genetically resistant chickens to meet the global protein consumption demand. Moreover, biopharming offers an excellent method to improve the quality of agriculture and livestock products ^[2]. Primordial germ cells (PGCs) are promising genetic resources for avian studies, including modified animals. Germ cells are known to transmit information to the upcoming generation with the help of gametogenesis. Gametogenesis is further described in this entreviewy. During development, PGCs are the first germ cell population. They are established and are the main origin of both oocytes and spermatogonia ^[3]. Chicken PGCs are model systems to understand avian and other animals’ gene functions ^[4]. Previous data have highlighted that PGC migration in all species has similar pathways/mechanisms. Lipids and G-protein-coupled receptors play an essential role in the regulation of PGCs ^[5]. PGCs are the homogeneous ancestors of gametes. These cells are responsible for transforming genetic data from one cohort to another ^[6]. In contrast to mammalian transgenic and genome-editing systems, transgenesis and genome editing in chickens and other birds only depend on a special system called germline transmission, involving PGCs. Animal phenotypes, genotypes, and traits are now easily modifiable owing to improvements in precise genome-editing technologies and genetic modification tools ^[7]. Animal breeders have historically used specific or unnatural breeding techniques to enhance the quality of food, productivity, and many other traits of offspring through the careful breeding of exceptional parents ^[8]. This method of discriminatory breeding is in line with the outcomes of contemporary genetic engineering or genome editing in terms of the intended genomic sequence of the animal’s DNA. Therefore, it has become possible to create more efficient traits with greater precision ^[9].

2. The Application of PGCs

2.1. Cryopreservation of PGCs—A Frozen Storage Technique

Over the past few decades, chicken breeds have faced various genetic changes; the evolution in their behavior, adaptation of traits, reproduction, and morphology are the main changes that they have faced during the past few decades. All of these things make their local varieties precious in terms of cultural heritage. The chicken is also considered to be the most numerous domestic animal. Seventy years ago, cryopreservation techniques were introduced to preserve specific specimens (i.e., semen and eggs). The process involves the collection of semen or eggs, evaluating them by dilution, cooling, equilibration, adding CPA, and then freezing in liquid nitrogen. Whenever the samples were subjected to usage again, a process of thawing was followed. Although cryopreservation is an excellent technique, it still contains many drawbacks. The significant steps of cryopreservation may lead to irregular cell structure, solution effects, membrane protein and lipid recognition, and oxidative stress. Cryopreservation is a method for preserving cells and tissues by using low temperatures. The cryopreservation of PGCs is performed via slow freezing or vitrification. Many studies have been conducted on the improvement of this technique. So far, weit is known that PGCs are valuable for cell-based genetic engineering, genetic modification and preservation, and germplasm expansion. Certainly, PGCs can be developed in culture as mentioned above and cryopreserved with alterations in their biological properties. As PGCs are vital for chicken transgenesis, their preservation is also essential. Consequently, PGCs appear to be the best candidates for cryopreservation. Two main methods are used for the cryopreservation of living cells and tissues: vitrification, and slow freezing. Slow freezing of PGCs is usually performed using serum-containing media with the addition of DMSO and ethylene glycol. Today, commercially prepared premixed media are also available, which can be used successfully. At the same time, less data and research are available for the cryopreservation of PGCs using the vitrification technique. However, vitrification is considered to be more efficient than slow freezing—especially for the cryopreservation of oocytes, stem cells, and embryos—in terms of cell survival and stability [60]^[10]. The significance of cryopreservation is mentioned briefly in this section. In short, sustainability and biosecurity in poultry production are challenging compared to other livestock species. This is because embryo preservation is not very feasible for egg-laying species. Therefore, cryopreservation of PGCs is a potential solution. This can help to produce sufficient independent lines of male and female PGCs that would be appropriate to rebuild a chicken breed. Cryopreservation of PGCs is a major step in biobanking. By using cryopreservation, we can conserve beneficial traits can be conserved to meet chickens’ current and future production needs.

2.2. Research Progress of Primordial Germ Cells (PGCs)—A Cell-Based Gene Transfer Method

In avian species, oocytes are the main precursor cells, commonly known as primordial germ cells. They are referred to as the inherited mode, as well as the preformation mode. There are a lot of gene transfer methods. Compared to other gene transfer methods, the cell-based gene transfer method is the most attractive. The cell-based approach to gene insertion is appealing because it enables the generation of transgenes with specific genome modifications and allows for the examination of transgenes’ expression and integration into the genome before cell implantation into the embryo. Obtaining cultures of cells that can reach the germline (i.e., primordial germ cells), cultivating them, transfecting them with DNA constructs, and implanting the transfected cells into an embryo constitute the initial stages in cell-based transgenic manufacturing. The G0 progeny are then mated to create G1 hens, which have the transgene implanted throughout their entire body. This technical framework combines the efforts of three technologies: the ability to cultivate germline-capable cells, the creation of acceptable DNA constructs, and the production of germline chimeras. The chicken germline chimera was first created by researchers in the lab [58]^[11]. The creation of avian germline chimeras has since become a routine process, and techniques for cultivating chicken germ cells, embryonic stem cells, and DNA vectors have all been established. Therefore, while in vitro germline alteration may be used to produce transgenic birds, it has not yet proven successful. Despite this, all of the necessary techniques are accessible. Finally, there have been some recent developments in sperm-mediated gene transfer. In a nutshell, sperm and a DNA construct are combined, and the combination is utilized to inseminate a fertile female. The sperm that efficiently fertilizes the egg carries the transgene of interest, which is then present in the resulting embryo and hatchlings. Transgenic mammal production has recently been claimed to be reproducible using sperm-mediated transfer. However, the successful integration of a transgene of interest into the avian germline has not yet been demonstrated using sperm-mediated gene transfer. To create transgenic birds, only retroviral and microinjection technology has been used to date [26,61]^[12][13].