BAC mediated transgenic Large White boars with FSHα/β genes from Chinese Erhualian pigs

Abstract

Follicle-stimulating hormone (FSH) is a critical hormone regulating reproduction in mammals. Transgenic mice show that overexpression of FSH can improve female fecundity. Using a bacterial artificial chromosome (BAC) system and somatic cell nuclear transfer, we herein generated 67 Large White transgenic (TG) boars harboring FSHα/β genes from Chinese Erhualian pigs, the most prolific breed in the world. We selected two F0 TG boars for further breeding and conducted molecular characterization and biosafety assessment for F1 boars. We showed that 8-9 copies of exogenous FSHα and 5-6 copies of exogenous FSHβ were integrated into the genome of transgenic pigs. The inheritance of exogenous genes conforms to the Mendel's law of segregation. TG boars had higher levels of serum FSH, FSHα mRNA in multiple tissues, FSHβ protein in pituitary and more germ cells per seminiferous tubule compared with their wild-type half sibs without any reproductive defects. Analysis of growth curve, hematological and biochemical parameters and histopathology illustrated that TG boars grew healthily and normally. By applying 16S rRNA gene sequencing, we demonstrated that exogenous genes had no impact on the bacterial community structures of pig guts. Moreover, foreign gene drift did not occur as verified by horizontal gene transfer. Our findings indicate that overexpression of FSH could improve spermatogenesis ability of boars. This work provides insight into the effect of FSHα/β genes on male reproductive performance on pigs by a BAC-mediated transgenic approach.

Gain–of–Function Genetic Models to Study FSH Action

“…There was no significant difference between semen quality parameters of Tg and WT boars. However, the number of germ cells per seminiferous tubule was significantly higher in Tg boars than WT (44). The elevated germ cell counts in seminiferous tubules suggested increased spermatogenesis capacity, but the lack of significant results from semen evaluation leaves room for further analysis to confirm this possibility (44).…”

“…Porcine FSH was further analyzed by the same group that produced BAC pFSH subunits from Chinese Erhualian pigs, and introduced them into Large White Boars (44). As the Large White Boar previously showed poor reproductive performance, the investigators sought to determine if overexpression of pFSH from a highly prolific pig breed could improve fertility.…”

“…Expression of FSHβ was localized specifically to the pituitary and was significantly higher in Tg than WT boars (44). Higher serum levels of both FSHα and FSHβ were observed in Tg boars suggesting overexpression of pFSH.…”

“…Higher serum levels of both FSHα and FSHβ were observed in Tg boars suggesting overexpression of pFSH. Male reproductive performance was measured by evaluating semen volume, sperm motility, sperm concentration, and total sperm number per ejaculation (44). There was no significant difference between semen quality parameters of Tg and WT boars.…”

“…In a third study done using the same BAC system containing pFSH α and β, analysis of reproductive phenotypes in female transgenic (Tg) Large White Boars were analyzed. Methods for pFSH BAC transfer were the same as previously described for this model (44). The specific integration site of pFSH into Large White Boars was determined using whole-genome sequencing, identifying exogenous FSHα/β genes at 140, 646, 456 bp on chromosome 9 (46).…”