The production of therapeutic protein from transgenic domestic animal is the major technology of biotechnology. Insulin-like growth factor-1 (IGF-1) is known to play an important role in the growth of the animal. The objective of this study is construction of knock-in vector that bovine IGF-1 gene is inserted into the exon 7 locus of β-casein gene and expressed using the gene regulatory DNA sequence of bovine β-casein gene. The knock-in vector consists of 5’ arm region (1.02 kb), bIGF-1 cDNA, CMV-EGFP, and 3’ arm region (1.81 kb). To express bIGF-1 gene as transgene, the F2A sequence was fused to the 5’ terminal of bIGF-1 gene and inserted into exon 7 of the β-casein gene. As a result, the knock-in vector is confirmed that the amino acids are synthesized without termination from the β-casein exon 7 region to the bIGF-1 gene by DNA sequence. These knock-in vectors may help to create transgenic dairy cattle expressing bovine bIGF-1 protein in the mammary gland via the expression system of the bovine β-casein gene.

Recently, with the increase of meat production, high quality and safety of meat have been strongly emphasized by Korean consumers. Marbling in beef has been regarded as an important criterion deciding meat quality in Korea. The purpose of this study was to identify the transcriptional level of insulin-like growth factor-1 (IGF-1) in longissimus muscle samples of 46 Hanwoo. The level of IGF-1 transcripts was measured by real-time polymerase chain reaction (PCR) and molecular connection of IGF-1 was analyzed using the Pathway Studio program (Ver 9.0). Increase of marbling score (MS) induced increase of IGF-1 transcripts level in the muscle and there is a significant correlation (p<0.05) between IGF-1 mRNA expression and MS. The pathway study showed that IGF-1 genes are regulated in insulin, fatty acid synthase, leptin, and corticotrophin releasing hormone. These results suggest that IGF-1 might be used as a useful marker for the improvement of economic traits in Hanwoo.

Differential capacity of the parthenogenetic embryonic stem cells (PESCs) is still under controversy and the mechanisms of its neural induction are yet poorly understood. Here we demonstrated neural lineage induction of PESCs by addition of insulin-like growth factor-2 (Igf2), which is an important factor for embryo organ development and a paternally expressed imprinting gene. Murine PESCs were aggregated to embryoid bodies (EBs) by suspension culture under the leukemia inhibitory factor-free condition for 4 days. To test the effect of exogenous Igf2, 30 ng/ml of Igf2 was supplemented to EBs induction medium. Then neural induction was carried out with serum-free medium containing insulin, transferrin, selenium, and fibronectin complex (ITSFn) for 12 days. Normal murine embryonic stem cells derived from fertilized embryos (ESCs) were used as the control group. Neural potential of differentiated PESCs and ESCs were analyzed by immunofluorescent labeling and real-time PCR assay (Nestin, neural progenitor marker; Tuj1, neuronal cell marker; GFAP, glial cell marker). The differentiated cells from both ESC and PESC showed heterogeneous population of Nestin, Tuj1, and GFAP positive cells. In terms of the level of gene expression, PESC showed 4 times higher level of GFAP expression than ESCs. After exposure to Igf2, the expression level of GFAP decreased both in derivatives of PESCs and ESCs. Interestingly, the expression level of Tuj1 increased only in ESCs, not in PESCs. The results show that IGF2 is a positive effector for suppressing over-expressed glial differentiation during neural induction of PESCs and for promoting neuronal differentiation of ESCs, while exogenous Igf2 could not accelerate the neuronal differentiation of PESCs. Although exogenous Igf2 promotes neuronal differentiation of normal ESCs, expression of endogenous Igf2 may be critical for initiating neuronal differentiation of pluripotent stem cells. The findings may contribute to understanding of the relationship between imprinting mechanism and neural differentiation and its application to neural tissue repair in the future.

Agi ng adversely affects the structure and function of the saliva ry gla nd and leads to ma rked insulin resis t ance that correlates with red uced ins ulin s ignal t ra nsduction, lnsulin-like g rowth factors are k.nown to be regulator involved in embryonic a nd postnatal development with sequences 62% identical t o that of proins ulin , To determine whether i n suli n- like growth facto l' - l, -2, - 1 receptor a l'e involved in the changes in rat salival'y gland by aging, we a na lyzecl the qua nti tation of 19f - 1, -2, - 1 rece ptor mRNA in l'at salivary glancl between 2 weeks and 26 months after bir th using the competiti ve reverse t ra nscriptase-polymerase chain reaction(RT-PCR) methocl Between 2 weeks ancl 2 months age, sharp falls in the qua nt ities of Igf-2 mRNA were observed, W11ereas Igf- 1 receptor mRNA rose by aging, but not sig nifïcantly, The quanti t ies of 19f- l kept by aging, These change seem to be involvecl a role fol' the Igf-2 in sali va ry c1evelopment a ncl earl y growth is incli ca ted, Thus, t he dras tic changes in the qua ntities of Igf-2 mRNA in the ra t sali va ry gla nd by aging seem to be in volved in the development, early growth and homeostasis of sali vary gla nd,

To deterrnine the effects of insulin-like growth factor system in the aging process of the rat submandibular gland, expression pattems of 19f-1, 19f-2 and 19f-1 receptor were investigated by immunohistochem엽따. 1n addition, age-related changes of 19f-1 and 19f-2 concentrations in serum were deterrnined by radioimmunoassay 19f-1, 19f-2 and 19f-1 receptor were principally expressed 띠 the duct cells. 19f-1 receptors were strongly expre잃ed in 2-week-old group but were gradually decreased with advancing age. 19f-1 expressions were the strongest in the excretory and striated ducts of 7-month-old and 17-month-old groups. 19f-1 expressions, however, were weakiy expressed in the cells of granular convoluted tubules of 7-month잉ld ， 17-month-old and 27-month-old groups. 19f-2 expressions were found in some cells around duαs of 2-week-old group but were persisted relatively weak. The concentrations of 19f-1 and 1향2 in serum were the highest in 2-week-old group but were decreased and maintained at low level with advancing age. These results show that all components of 19f system were expressed in the rat submandibular giand in an age-related manner. Serum concentrations of all components of 19f system were not dosely related with the intra-밍and비ar expresslon patterns 까lerefore ， 1양 systerns may play important roles in the aging process of the rat submandibular gland in an autocrine/paracrine manner

본 연구는 돼지 체외수정란의 배양시스템의 최적화를 위하여 수행되었다. 기본배양액으로는 NC-SU-23을 사용하였으며, 실험 1에서는 IGF-I이 각각 0, 1, 10, 100 ng/의 농도로 첨가된 배양액에서 배양하여 분할율, 배반포 발달율과총 세포수를 확인하였다. 농도 경사에 따른 분할율은 차이가 없었으나(65.5%~69.5%), 배반포 발달율에 있어서는 10 ng/의 농도로 첨가하였을 경우가 가장 높았다(P<0.05). 실험 2는 배 아 유전자 활성

Epidermal growth factor (EGF) and insulin-like growth factor-I (IGF-I) have been shown to stimulate proliferation and differentiation of various somatic cells, including placental trophoblasts and also to enhance fetal growth and development when maternally administered. Since an increase of the expression of placental EGF and IGF-I receptors in rat, mouse, and human with the gestation advanced, both EGF and IGF-I were considered to play pivotal roles on fetal growth by regulating some function of placental cells. Amino acids are crucial importance for both maternal and fetal requirements of energy source and essential constituent of fetal mass during pregnancy. Impaired fetal and placental uptake of amino acids has been observed in several models of growth retardation in the rat. Amino acid is concentrated in the fetal side through active transport by amino acid transporters and is one of the important metabolic fuels for the fatal growth. Therefore, at first plasma amino acid concentrations in mothers and fetuses were measured as an index of uphill transport across the placenta associated with EGF and IGF-1. The EGF administration at the concentration of 0, 0.1, or 0.2 /g to pregnant rats from day 18 to 21 of gestation apparently increased fetal/maternal ratio of serum proline concentration and also fatal growth in EGF dose-dependent manner. When IGF-I in doses of 0, 1, 2, and 4 /g were administrated, the ratio of leucine, isoleucine, tryptophan, phenylalanine, tyrosine and also fetal growth significantly increased with a dose-dependent manner. These results suggested that EGF and IGF-I enhanced fatal growth by, as one of its possible mechanisms, promoting placental activity to transfer some amino acid supplies from the mother to the fetus in late pregnancy.