• Korean Journal of Medicinal Crop Science
• /
• v.15 no.5
• /
• pp.339-345
• /
• 2007

Field performance and morphological characterization was conducted on seven transgenic lines of Codonopsis lanceolata expressing ${\gamma}-TMT$ gene. The shoots were obtained from leaf explants after co-cultivation with Agrobacterium tume-faciens strain LBA 4404 harboring a binary vector pYBI 121 that carried genes encoding ${\gamma}-Tocopherol$ methyltransferase gene (${\gamma}-TMT$) and a neomycin phosphotransferase II gene (npt II) for kanamycin resistance. The transgenic plants were transferred to a green house for acclimation. Integration of T-DNA into the $T_0\;and\;T_1$ generation of transgenic Codonopsis lanceolata genome was confirmed by the polymerase chain reaction and southern blot analysis. The progenies of transgenic plants showed phenotypic differences within the different lines and with relative to control plants. When grown in field, the transgenic plants in general exhibited increased fertility, significant improvement in the shoot weight, root weight, shoot height and rachis length with relation to the control plants. However, all seven independently derived transgenic lines produced normal flower with respect to its shape, size, color and seeds number at its maturity. Indicating that the addition of a selectable marker gene in the plant genome does not effect on seed germination and agronomic performance of transgenic Codonopsis lanceolata. $T_1$ progenies of these plants were obtained and evaluated together with control plant in a field experiment. Overall, the agronomic performance of $T_1$ progenies of transgenic Codonopsis lanceolata showed superior to that of the seed derived non-transgenic plant. In this study, we report on the morphological variation and agronomic performance of transgenic Codonopsis lanceolata developed by Agrobacterium transformation.

• Korean Journal of Veterinary Service
• /
• v.18 no.2
• /
• pp.113-123
• /
• 1995

The present study was conducted to investigate the prevalent characteristics of Fowl Typhoid and antibiotic susceptibility of Salmonella gallinarum isolated from 56 infective or dead chickens of 20 egg laying farms in Kyung Buk province during the period from August to December 1994. 1. Among 416, 000 chickens of 92 flocks in 20 egg laying farms, 17, 360 chickens of 31 flocks were died of Fowl Typhoid. 2. Salmonella gallinarum was isolated from 56 chickens in liver and spleen, and then blood of infective chickens was positive to Pullorum antigen. 3, In the survey of gross lesion of 56 chickens, 43 chickens(76.8%) were swelled at liver, 39(69.6%) were swelled at spleen, 12(21.4%) were changed with bronze, 3(5.4% ) were hemorrhagic in peritoneal cavity. 4. In transmission pattern, 4 farms were outbreaked the entrance of chicken house at first, but the others were outbreaked at various place. They were transmitted at right and left directions in flock. 5. 2 farms confirmed at the early stage of infection were eradicated by removing infective chickens and administrating antibiotics, but 18 farms at chronic stage were not. 6. The biochemical properties of 112 Salmonella gallinarum from chickens were generally identical to those of the referance, but H$_2$S was not productive, cellobiose was fermentive. 7 Minimum inhibitory concentration(MIC) of 20 isolates was performed by using 21 antibiotics, MICs of Amikacin(Ak), Gentamicin(Gm), Kanamycin(Km), and Tetracycline (Tc) were below 1.6 ug/ml, Ampicillin(Am), Furazolidone(Fu) and Neomycin(Nm) were below 3.1 ug/ml, Cephalothin(Ce), Cefazoline(Cf) and Chloramphenicol(Cm) were below 6.3 ug/ml, Nalidixic acid(Na), Polymyxin(Po) and Rifampicin(Rf) were below 12.5 ug/ml, Penicillin (Pm) was below 25 ug/ml, Colistin(Co) and Streptomycin(Sm) were below 50 ug/ml, Sulfamerazine(Sr) and Sulfamethazine (St) were below 200 ug/ml, Lincomycin(Lm) and Spiramycin(Sp) were below 400 ug/ml, Bacitracin(Ba) was below 800 ug/ml. 8. Among the 20 isolates, all(100%) of those were sensitive to Ak, Am, Ce, Cf, Cm, Fu, Gm, Km, Na, Nm, Po, Rf, Sr, St and Tc, but 6 isolates(30%) were resistent to Co, 20(100% ) to Ba, Lm, Pm, Sm, and Sp. The drug resistance patterns were simple which 6 strains were BaCoLmPmSmSp type, and 14 were BaLmPmSmSp type.

• Reproductive and Developmental Biology
• /
• v.32 no.3
• /
• pp.205-209
• /
• 2008

This study was carried out to develop knock-in vector for EGFP (enhanced green fluorescent protein) expression in porcine $\beta$-casein locus. For construction of knock-in vector using porcine $\beta$-casein gene, we cloned the $\beta$-casein genome DNA from porcine fetal fibroblast cells, EGFP and SV40 polyA signal using PCR. The knock-in vectors consisted of a 5-kb fragment as the 5' recombination arm and a 2.7-kb fragment as the 3' recombination arm. We used the neomycin resistance gene ($neo^{r}$) as a positive selectable marker and the diphtheria toxin A (DT-A) gene as a negative selectable marker. To demonstrate EGFP expression from knock-in vector, we are transfected knock-in vector that has EGFP gene in murine mammary epithelial cell line HC11 cells with pSV2 neo plasmid. The EGFP expression was detected in HC11 cells transfected knock-in vector. This result demonstrates that this knock-in vector may be used for the development of knock-in transgenic pig.

• Reproductive and Developmental Biology
• /
• v.41 no.1
• /
• pp.7-16
• /
• 2017

The knock-in efficiency in the fibroblast is very important to produce transgenic domestic animal using nuclear transfer. In this research, we constructed three kinds of different knock-in vectors to study the efficiency of knock-in depending on structure of knock-in vector with different size of homologous arm on the ${\beta}-casein$ gene locus in the somatic cells; DT-A_cEndo Knock-in vector, DT-A_tEndo Knock-in vector I, and DT-A_tEndo Knock-in vector II. The knock-in vector consists of 4.8 kb or 1.06 kb of 5' arm region and 1.8 kb or 0.64 kb of 3' arm region, and neomycin resistance gene(neor) as a positive selection marker gene. The cEndo Knock-in vector had 4.8 kb and 1.8 kb homologous arm. The tEndo Knock-in vector I had 1.06 kb and 0.64 kb homologous arm and tEndo Knock-in vector II had 1.06 kb and 1.8 kb homologous arm. To express endostatin gene as transgene, the F2A sequence was fused to the 5' terminal of endostatin gene and inserted into exon 7 of the ${\beta}-casein$ gene. The knock-in vector and TALEN were introduced into the bovine fibroblast by electroporation. The knock-in efficiencies of cEndo, tEndo I, and tEndo II vector were 4.6%, 2.2% and 4.8%, respectively. These results indicated that size of 3' arm in the knock-in vector is important for TALEN-mediated homologous recombination in the fibroblast. In conclusion, our knock-in system may help to create transgenic dairy cattle expressing human endostatin protein via the endogenous expression system of the bovine ${\beta}-casein$ gene in the mammary gland.

• The Journal of the Korean Society for Microbiology
• /
• v.21 no.1
• /
• pp.73-95
• /
• 1986

A total of 5,462 isolates suspicious of Salmonella, Shigella and E. coli which were isolated during 1983 to 1985 by 12 City Hygine Laboratories and General Hospital Laboratories were received and identified at the National Salmonella Center, Seoul, Korea. The result of identification of these strains were summarized as follows: 1. It was confirmed that the total organisms broke down into 2,014 strains of Salmonella 1,294 of which were S. typhi, 887 strains of Shigella and 2,561 strains of E. coli. 2. For seasonal distribution of enteric pathogens, July was the month with the highest out breaks of salmonellosis, May was the month of Shigellosis, and April was of the highest month it in the case of E. coli. 3. Salmonella typhi with the highest incidence of isolation was shown to belong to various phage types, especially with the strains detected in Seoul. M1 type was widely distributed all over the country, but the majority was E1 type in 1983. 4. For age distribution of patients, the 20-29 age group had the highest incidence of salmonellosis whileas the 1 to 9 age group had the highest incidence of Shigellosis. 5. For sexuly distribution of Salmonella and Shigella infections seemed to be relatively higher in the female than in the male. However, E. coli. had no relationship to both sex. 6. The antibiotic sensitivity patterns of S. typhi cultures showed a tendancy to be resistant to colistin, gentamycin, neomycin, tetracycline and streptomycin. 7. The isolates of S. paratyphi-A, S. typhimurium and S. enteritidis seemed to have a tendency of multiple drug resistance. 8. 93.9 percent of 1,568 E. coli strains showed negative reactions to the antisera of enteropathogenic E. coli and 15.6 percent of them produced a heat-labile enterotoxin, but positive reaction to the antisera was 6.1 percent and 11.6 percent of them producled the enterotoxin.

• Current Research on Agriculture and Life Sciences
• /
• v.2
• /
• pp.115-121
• /
• 1984

Some investigations on chronic mastitis in dairy cattle in Taegu-Kyungpook Provinces from the beginning of October, 1984 till the end of August, 1985 were conducted with the particular regard to the causative agents and their drug susceptibility. Milk samples from 83 isolated cases of chronic mastitis cattle were investigated bacteriologically and the causative organisms recovered were examined for their antibiotic susceptibility by using disc diffusion susceptibility technique against the major antibiotics of current veterinary use. Major causative agents involved in chronic mastitis in Taegu-Kyungpook Provinces were in order of prevalence Staphylococcus spp. (48.2 %), Escherichia coli (18.1 %), Candida spp. (10.8 %) and Corynebacterium spp. (8.4 %), Streptococcus agalactiae (3.6 %), Bacillus cereus (3.6 %) and Pseudomonas aeruginosa (2.4 %) were found to be one of the minor agents. The majority of staphylococcal isolates and E. coli were highly resistant to the most antibiotics tested. The percentages of staphylococcal cultures resistant to penicillin, methicillin, lincomycin, novobiocin, ampicillin and tetracycline were 87.2 %, 78.7 %, 68.1 %, 61.7% and 57.4 %, respectively, while the majority of them were susceptible to gentamicin(78.7 %), cephalothin(76.6 %) and chloramphenicol (74.5%). E. coli isolates were found to be highly resistant to streptomycin, cephalothin, tetracycline and ampicillin while the majority of them were susceptible to colistin (83.3 %), gentamicin (77.8 %) and chloramphenicol (66.7 %). Corynebacterium spp. were susceptible to ampicillin, chloramphenicol, erythromycin, gentamicin, oleandomycin and tetracycline although they showed resistance to novobiocin and penicillin. Two cultures of Pseudomonas aeruginosa recovered from mastitis milk were highly resistant to the antibiotics employed in the present study.

• Korean Journal of Animal Reproduction
• /
• v.21 no.2
• /
• pp.167-176
• /
• 1997

This study was carried out to evaluate the potential for preselection of transgenic embryos prior to transfer into recipient animals. In these experiments, I used a 3.2 kb transgene which contained the neomycin resistance gene (neo) and lac Z gene driven by the $\beta$ actin promoter (iacZ Ineo). Oocytes were aspirated from abattoir ovaries, matured in TCM-199 supplemented with 10% fetal bovine serum (FBS), 5 ${\mu}\textrm{g}$/ml LH, 0.5 ${\mu}\textrm{g}$/ml FSH, 100 unit/ml penicillin, and 100 ${\mu}\textrm{g}$/ml streptomycin for 22 to 24 hrs then inseminated with a sperm suspension of 1 X 10$^6$ sperm/ml containing 5 ${\mu}\textrm{g}$/ml of heparin. At 18-20 hrs after insemination, cumulus cells were removed by vortexing and pronuclei of centrifuged zygotes microinjected with the lacZ/neo construct (3 ng/$\mu$l). All cultures were carried out in CR1aa with transfected BRL monolayers containing 3 mg/ml BSA, 20 $\mu$/ml NEM amino acids and 40 $\mu$I/ml BME amino acids. To identify the appropriate concentration of G418 for selection, non-microinjected zygotes were cultured in the presence of 0, 50, 100 and 200 $\mu$g/ml of G418. After 8 days of culture in these treatments, 44/145 (30.3%), 13/150 (8. 7%), 1/151 (0.7%) and 0/134 (0.0%) devel-oped to the blastocyst stage in 0, 50, 100 and 200 $\mu$g/ml of G418, respectively. A total of 1,127 zygotes were microinjected and placed into culture (without G418) and subsequently 710 (63.0%) cleaved. At 48 hrs post-injection, embryos ($\geq$2-cell) were randomly assigned to control (medium alone) or G418 (100 ${\mu}\textrm{g}$/ml) treatments. A control culture of 740 non-microinjected embryos from the same replicates of embryos were also placed into control medium. After 8 days in culture, 54/343 (15.7%) and 22/367 (6.0 %) of the microinjected embryos developed to the blastocyst stage in control and G418 media, respectively. A total of 151/740 (27.2%) of the non-microinjected embryos placed in the control medium developed to the blastocyst stage. The blastocysts in the control treatment had a mean of 70.7 ${\pm}$ 4.7 cells of which 23.1 ${\pm}$ 2.6 (32.7%) stained for $\beta$-Gal activity. B1astocysts in the G418 treatment had a mean of 48.8${\pm}$7.5 cells of which 40.3 ${\pm}$ 4.1 (82.6%) stained for $\beta$-Gal ac tivity (P<0.05). In the control treatment 26 of 30 (87.0%) blastocysts had some cells with $\beta$-Gal activity while all of the blastocysts in the G418 treatment had some cell with $\beta$-Gal ac tivity (P<0.05). However, ICM colonies in either control or G418 treatments were not expressed any epiblast cell except of trophetoderm celIs. The $\beta$-actin promoter/lacZ gene may not be e expression or silence expression in epiblast cells These results clearly show an enrichment of blastocysts expressing the transgene in the majority of their cells after culture in the presence of G418. The exogeneous gene was not express a and silence in ICM colonies especiallly epiblast cells except of trophectederm cells. Even though the higher rate cell number expressed of exogeneous gene in the G418 treatments, a total cell number was decrease significantly (P<0.05) of which might be also drop of the establishment of ES like-cell colonies and production of transgenic animals. However, futher studies need to determine the viability of these selected embryos and the avability of production of transgenic offspring.