Genetic screening is the prominent application of haploid stem cell technology. Saccharomyces cerevisiae was used as a model organism to study haploid gene function before the derivation of haESCs [1]. However, due to species specificity, this approach cannot be directly applied to mammalian systems. Although the human genome project has successfully sequenced all 3 billion chemical units and identified approximately 20,000–25,000 genes in the human genome, their relevant functions are yet to be elucidated fully and require further study. Global genomic screening has therefore been widely used in mammalian ESCs in order to clarify gene function in many biological procedures. However, it is difficult to obtain homozygous mutations in diploid ESCs. Additionally, heterozygous genotypes may have no impact on their phenotype for the homologous allele complement. This means that diploid genomes hamper the study of recessive genetic conditions. Since haploid cells only carry one set of chromosomes, they exhibit corresponding phenotypes in the presence of a mutation. Genetic screening in mammalian cells often directly promotes medicinal and pharmacological research. Currently, it is difficult to obtain double homogeneous allele mutations through genome editing techniques. This hinders the generation of homozygous gene knockout libraries. The use of haESCs, however, can overcome this obstacle.

Typically, genetic screening aims to obtain loss-of-function phenotypes through allele mutation. Genomic engineering is applied to produce mutant libraries through transposon-mediated insertion or nuclease-mediated targeting modification technologies—including

piggyBac

, Clustered regularly interspaced short palindromic repeats/Cas (CRISPR/Cas), and transcription activator-like (TAL) effector nucleases (TALENs) [

34

]. Once screened and tested, specific mutated cells are said to have been generated. Hitherto, through gene screening on haploid cell lines, basic studies including DNA repair, drug toxicity, X-chromosome inactivation, cell differentiation, and human clinical diseases have been investigated separately [

3

5

,

7

,

8

,

23

,

29

,

35

43

] (Fig. 

3

). Primate haESCs can maintain haploidy in long-term culture, which makes them good resources for gene function research [

5

,

7

,

8

]. All of the established mammalian haploid cells, including near-haploid human tumor cells, have been applied in whole-genome genetic screening at the cellular or organism level, and many accomplishments have been achieved to date. Leeb et al. [

37

] found that Zfp706 and Pum1 were key regulators governing differentiation of naïve stem cells by screening mutated haESCs. Recent gene editing techniques facilitated the efficient utilization of haESC resources, such as CRISPR/Cas9 knockout libraries, to acquire offspring carrying multiple heterozygous mutations [

23

,

44

]. In addition, gene trapping with a

piggyBac

transposon is a more efficient approach in the genetic screening of haESCs, because it allows more precise assessment of the integration site than when using CRISPR/Cas or TALENs [

6

,

45

]. Taken together, gene screening in haESCs boosts the basic and clinical research fields of developmental biology and regenerative medicine [

34

]. This holds great value in the study of cancer, species evolution, biomolecular interactions, lineage specification, and signal pathways.

Fig. 3

Application of haESCs in multiple types of genetic screening. Since haESCs contain a haploid copy of the genome, it is easy to obtain homozygous mutant libraries through transposon or viral systems. To date, mutant haESC libraries have been exploited in various targeted gene screening systems in order to track cell fate. Such systems include factors such as toxic resistance [4, 5], cell differentiation [6, 37, 48], and X-chromosome inactivation [7, 35, 36]. haESC haploid embryonic stem cell