• Asian-Australasian Journal of Animal Sciences
• /
• 제25권9호
• /
• pp.1223-1228
• /
• 2012

The population sizes of three Korean indigenous cattle populations have been drastically reduced over the past decades. In this study, we examined the extent to which reduction in populations influenced genetic diversity, population structure and demographic history using complete mitochondrial DNA (mtDNA) control region sequences. The complete mtDNA control region was sequenced in 56 individuals from Korean Black (KB), Jeju Black (JEB) and Korean Brindle (BRI) cattle populations. We included 27 mtDNA sequences of Korean Brown (BRO) from the GenBank database. Haplotype diversity estimate for the total population was high (0.870) while nucleotide diversity was low (0.004). The KB showed considerably low nucleotide (${\pi}$ = 0.001) and haplotype (h = 0.368) diversities. Analysis of molecular variance revealed a low level of genetic differentiation but this was highly significant (p<0.001) among the cattle populations. Of the total genetic diversity, 7.6% was attributable to among cattle populations diversity and the rest (92.4%) to differences within populations. The mismatch distribution analysis and neutrality tests revealed that KB population was in genetic equilibrium or decline. Indeed, unless an appropriate breeding management practice is developed, inbreeding and genetic drift will further impoverish genetic diversity of these cattle populations. Rational breed development and conservation strategy is needed to safeguard these cattle population.

• Asian-Australasian Journal of Animal Sciences
• /
• 제25권7호
• /
• pp.921-926
• /
• 2012

In order to protect the genetic resource of native horse breeds, the genetic diversity of mitochondrial DNA (mtDNA) D-loop of three native horse breeds in western China were investigated. Forty-three 600 bp mtDNA D-loop sequences were analyzed by PCR and sequencing techniques, 33 unique haplotypes with 70 polymorphic sites were detected in these horses, which account for 11.67% of 600 bp sequence analyzed, showing the abundant genetic diversity of the three native horse breeds in western China. The Neighbour-Joining (NJ) phylogenetic tree based on 247 bp of 43 D-loop sequences demonstrated the presence of seven major lineages (A to G), indicating that the three native horse breeds in western China originated from multiple maternal origins. Consistent with the front, the NJ phylogenetic tree based on 600 bp of mtDNA D-loop sequences of 43 Chinese western native horses and 81 sequences of six horse breeds from GenBank indicated that the three horse breeds had distributed into the seven major lineages (A to G). The structure of the phylogenic tree is often blurred because the variation in a short segment of the mitochondrial genome is often accompanied by high levels of recurrent mutations. Consequently, longer D-loop sequences are helpful in achieving a higher level of molecular resolution in horses.

• Asian-Australasian Journal of Animal Sciences
• /
• 제18권6호
• /
• pp.892-897
• /
• 2005

Macrophage colony-stimulating factor (M-CSF) is a growth factor required for growth and differentiation of mononuclear phagocyte lineage. Total and 16 poly (A) mRNA of bovine M-CSF were isolated from healthy bovine peripheral mononuclear cells stimulated by phobol 12-myristste 13-acetate (TPA). The more compatible cultured mononuclear cells were 5${\times}$10/ml for RNA isolation. TPA-activated mononuclear cells increased the level of M-CSF-mRNA more than concanavalin A (Con A) and lipopolysaccharide (LPS). The optimal analysis of reverse transcriptase-polymerase chain reaction (RT-PCR) for14 Macrophage colonystimulating factor (M-CSF) as a growth factor required for bovine M-CSF was denaturation at 94$^{\circ}C$ for 1 minute, annealing at 57$^{\circ}C$ for 1 minute, extension at 72$^{\circ}C$ for 1 minute for 30 cycles. The size of cDNA of bovine M-CSF by RT-PCR was 774 base pairs. A 774 base pairs cDNA encoding bovine M-CSF was synthesized by reverse transcriptase polymerase chain reaction (RT-PCR). Ligated cDNA was transformed to competent cells and then plasmid isolation and digestion was performed. Molecular cloning and sequencing were performed for cDNA of bovine M-CSF. The size of cloned cDNA of bovine M-CSF was 774base pairs. The homology of base sequence and amino acid sequence was 88% and 86% compared with known human M-CSF, respectively. From a high degree of sequence similarity, the obtained cDNA of bovine M-CSF is thought be a specific gene of bovine M-CSF.

• Asian-Australasian Journal of Animal Sciences
• /
• 제17권7호
• /
• pp.897-902
• /
• 2004

PSMC5 subunit, which belongs to the 26S proteasomal subunit family, plays an important role in the antigen presentation mediated by MHC class I molecular. Full-length cDNA of porcine PSMC5 was isolated using the in silico cloning and rapid amplification of cDNA ends (RACE). Amino acid was deduced and the primary structure was analyzed. Results revealed that the porcine PSMC5 gene shares the high degree of sequence similarity with its mammalian counterparts at both the nucleotide level and the amino acid level. The RT-PCR was performed to detect the porcine PSMC5 expression pattern in seven tissues and the result showed that high express level was observed in spleen, lung, marrow and liver while the low express level was in muscle. The full-length genomic DNA sequence of porcine PSMC5 gene was amplified by PCR and the genomic structure revealed that this gene was comprised by 12 exons and 11 introns. Best alignment of the cDNA and genomic exon DNA sequence presents 4 mismatches and this information potentially bears further study in gene polymorphisms.

• Asian-Australasian Journal of Animal Sciences
• /
• 제18권7호
• /
• pp.933-941
• /
• 2005

To develop a functional cDNA chip, specific genes expressed in the tissues of swine Kagoshima Berkshire were screened. A total of 4,434 ESTs were obtained by constructing a cDNA library from total RNA isolated from the muscle and fat tissues, affirming their functions by investigating similarity of nucleotide sequences with the database at the NCBI. Among them, 1,230 ESTs were confirmed as novel genes, which, to date, have not been identified. Attaching the genes to a cDNA microarray slide revealed expression patterns of genes in muscle and fat according to the growth stages of swine. As specific genes expressed in the muscle tissues of swine with body weight of 30 kg, 60 genes including actin, myosin, tropomysin, transfer RNA-trp synthetase, Kel-like protein 23, KIAA0182 and COI, Foocen-m, etc were obtained. In addition, 18 novel genes were obtained. As specific genes expressed in fat tissues of swine with body weight of 30 kg, 47 genes including annexin II, Collagen, Fibronectin, Pleckstrin homology domain, serine protease, etc were obtained. 21 novel genes were also obtained. The genes specifically expressed in the muscle and fat tissues of swine affect contraction and relaxation of the muscle and the fat. However, studies on the expression mechanisms of the genes are insufficient. To reveal species of structural genes in swine muscle and fat tissue, interrelation studies in expression and function of genes by using the cDNA chip should be conducted.

• 식물조직배양학회지
• /
• 제26권4호
• /
• pp.245-250
• /
• 1999

수분 후 담배의 배에 중이온 ($^{14}$ N) beam을 조사하여 유도한 variegated담배의 잎 조직을 기내배양하여 재생된 식물체의 특성을 조사하였다. NAA 0.1mg/L 와 BAP 1/0mg/L를 첨가한 MS배지에서 variegated담배의 잎을 배양하면 백색부위에서는 백색 식물체만이 유도되었으나 녹색부위에서는 녹색 식물체가 47.2%, 백색 식물체가 37.4%, variegated식물체가 15.4% 유도되었다. 재생된 녹색 식물체는 자가수분한 후대 (F$_1$)에서 녹색 식물체 1,651 개:백색 식물체 54개로 분리되었다. 또한 variegated 잎에서 재생된 녹색 식물체와 정상주를 정역교배 한 결과, 전부 녹색체가 분리되었다. 이 결과는 variegated잎의 표현형 유전은 최소한 모성 유전이 아님을 보여준다. Variegated잎의 녹색부위를 배양하여 재생된 variegated식물체의 잎을 이용하여 DNA gel blot한 결과, 엽록체 유전자인 psbA, rbcL, 165 rDNA, 23S rDNA의 양은 흰색부위 잎에서 정상주인 녹색 잎과 variegated식물체 잎의 녹색부위 보다 많았다.

• 생명과학회지
• /
• 제27권5호
• /
• pp.524-529
• /
• 2017

본 연구에서는 zebrafish에 인간의 KCNE1 유전자가 삽입된 형광단백질 vector를 microinjection하고, 그 발현 여부를 확인하고자 하였다. 먼저 양 말단에 제한효소(EcoRΙ, BamHΙ) site를 넣어 제작한 primer들로 genomic DNA에서 KCNE1 유전자를 분리하였다. 그 결과는 약 402 bp 크기의 DNA band였고 이 PCR 산물을 형광단백질 vector인 pPB-CMVp-EF1-GreenPuro 속에 클로닝하여 pPB-CMVp-hKCNE1-EF1-GreenPuro plasmid를 제작하였다. 이렇게 준비된 형광 vector를 zebrafish 수정란에 microinjection하였고, 부화된 치어에서 RT-PCR과 DNA sequencing을 통해 GFP 및 hKCNE1의 발현을 최종 확인하였다. 본 연구는 향후 QT 연장증후군(LQTs)에 대한 동물 모델로써 신경자극 전도, 유전자 치료, 유용 유전자 클로닝을 위한 기술 개발에 응용될 수 있을 것으로 기대된다.

• 미생물학회지
• /
• 제36권2호
• /
• pp.103-108
• /
• 2000

한국에서 분리된 전염성 조혈괴저바이러스(infectious hematopoietic necrosis virus, IHNV)인 IHNV-PRT의 non-viron(NV)단백질을 암호화하고 있는 cDNA를 클로닝하여 이들의 염기서열을 분석하였다. NV는 336bpzmrl의 open reading frame을 포함하였으며 이로부터 111개의 아미노산 서열을 외국에서 분리된 IHNV들과 비교 분석한 결과 90-95%의 상동성을 보였다. 이러한 사실은 INHV의 NV단백질 유전자들은 IHNV의 strain에 관계없이 매우 보존되어 있음을 나타내준다. Northern blotting을 사용하여 NV의 발현을 측정한 결과 감염 후 20 시간분터 발현이 증가함을 확인 힐수 있었다. NV가 바이러스의 증식에 필요한지의 여부를 확인하기 위하여 바이러스 유전자의 antisense DNA를 사용하여 바이러스 증식 억제에 관한 실험을 수행하였다. Glycoprotein (G)의 antisense DNA를 처리한 경우 바이러스의 증식이 거의 억제된 반면 NV에 대한 antisense DNA를 처리한 경우 바이러스 증식에 거의 변화가 없었다. 이로부터 배양중인 세포가 있어서 NV는 증식에 필수적이지 않은 것으로 판단된다.

• 한국자원식물학회지
• /
• 제16권3호
• /
• pp.238-244
• /
• 2003

더덕 뿌리에서 유래한 EST 라이브러리로부터 dehydrin 유전자와 높은 상동성 을 나타내는 full clone cDNA를 얻었다. 더덕의 dehydrin, ClDhn1은 893 bp의 cDNA로 159개의 아미노산을 코딩하는 480 bp의 ORF를 가지고 있다. ClDhn1의 아미노산을 분석해 보면, 전체적으로 높은 친수성을 나타내며, lysine이 풍부한 K 반복구간(KIKEKLPG)을 카르보닐기 쪽에 2개 가지고 있다. 또한, 여러 dehydrin들의 공통적인 특징인 7개의 연속적인 serine잔기가 첫 번째 K반복 구간 앞에 위치한다. 그러나, 아미노기쪽의 DEYGNP보존 구간은 변형(DEHGNP)되어 있다. ClDhn1 유전자는 전사 단계에서 더덕의 뿌리에서 가장 높은 발현 양상을 보이며, 줄기와 잎에서는 적은 양이 발현되었다.

• Asian-Australasian Journal of Animal Sciences
• /
• 제32권6호
• /
• pp.891-895
• /
• 2019

Objective: Aim of present study was the set up of a fast and reliable protocol using species-specific markers for the quali-quantitative analysis of DNA and the detection of ruminant biological components in dairy products. For this purpose, the promoter of the gene coding for the ${\alpha}$-lactoalbumin (LALBA) was chosen as possible candidate for the presence of short interspersed nuclear elements (SINEs). Methods: DNA was isolated from somatic cells of 120 individual milk samples of cattle (30), Mediterranean river buffalo (30), goat (30), and sheep (30) and the gene promoter region (about 600/700 bp) of LALBA (from about 600 bp upstream of exon 1) has been sequenced. For the development of a single polymerase chain reaction (PCR) protocol that allows the simultaneous identification of DNA from the four species of ruminants, the following internal primers pair were used: 5'-CACTGATCTTAAAGCTCAGGTT-3' (forward) and 5'-TCAGA GTAGGCCACAGAAG-3' (reverse). Results: Sequencing results of LALBA gene promoter region confirmed the presence of SINEs as monomorphic "within" and variable in size "among" the selected species. Amplicon lengths were 582 bp in cattle, 592 bp in buffalo, 655 in goat and 729 bp in sheep. PCR specificity was demonstrated by the detection of trace amounts of species-specific DNA from mixed sources ($0.25ng/{\mu}L$). Conclusion: We developed a rapid PCR protocol for the quali-quantitative analysis of DNA and the traceability of dairy products using a species-specific marker with only one pair of primers. Our results validate the proposed technique as a suitable tool for a simple and inexpensive (economic) detection of animal origin components in foodstuffs.