• 농업과학연구
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• 제28권2호
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• pp.92-98
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• 2001

락토페리신은 다양한 생리활성을 나타내는 락토페린(약 80kD)에서 유래된 항균성펩타이드 분획물(5kD)이다. 마우스HC11유선상피세포에서 인체 락토페리신의 발현은 bovine beta-casein을 promotor로 하고 인체 락토페리신 cDNA를 삽입하여 제작한 pBL1-cin발현벡타를 이용하였다. 이 발현벡타를 이용하여 인체 락토페리신 발현여부를 RT-PCR, northern blot, dot blot분석을 통하여 확인하였다. pBL1-cin 발현백타를 HC11세포에 transfection 하여 얻은 RNA를 이용하여 RT-PCR를 한 결과 150bp의 크기로 확인되었고 Northern blot 분석결과는 약 2.3 kb의 크기로 확인되었다. 인체 락토페린 polyclonal항체를 이용하여 dot blot한 결과 인체 락토페리신이 분비됨을 확인하였다.

• 식물조직배양학회지
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• 제28권2호
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• pp.87-90
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• 2001

배추 유래의 cytosolic glutathione reductase 유전자 (BcGR1)의 지속적 발현과 형질전환 식물체의 oxidative stress에 대한 내성과의 관계를 분석하기 위하여, BcGR1 유전자를 CaMV 35S promoter의 하류에 연결한 다음, 담배에 형질전환하였다. PCR 및 Southern blot 분석을 통하여 BcGR1 유전자가 정상적으로 삽입된 32 계통의 T$_{0}$ 식물체를 선발하였다. Northern blot 분석 결과, 도입된 유전자가 형질 전환 식물체 내에서 항상적으로 발현된다는 것을 확인하였으며, 도입 유전자의 copy number와 발현량 사이에는 정의 상관관계를 보이지 않았다.

• 한국초지조사료학회지
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• 제21권1호
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• pp.21-26
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• 2001

환경 스트레스에 의해 야기되는 활성 산소종에 의한 피해에 내성을 가지는 목초의 개발을 위하여 오차드그래스의 배반 조직 유래의 캘러스에 배추유래의 cytosolic glutathione reductase 유전자(BcGRl)를 Agrobucterium tumefaciens EHA101을 매개로 형질전환시켰다. Hygromcin으로 선발된 캘러스로부터 재분화된 식물체는 야생형과 비교하여 형태적으로 차이를 나타내지 않았다. PCR 및 Southern blot 분석을 통하여 형질전환 식물체의 염색체 내에 BcGRl 유전자가 integration 되었음을 확인하였다. 오차드그래스의 잎으로부터 total RNA를 분리하여 Northern blot 분석을 실시한 결과, 도입된 유전자가 형짙전환 식물체 내에서 지속적으로 발현된다는 것을 확인하였다.

• 한국가축번식학회지
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• 제18권2호
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• pp.133-139
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• 1994

Two human lactoferrin expression vectors(pCChcLf and pCChcLf-1) were constructed using rat $\beta$-casein gene and human lactoferrin cDNA. The recombinant DNAs containing human lactoferrin cDNA were microinjected into the fertilized eggs of hybrid mice (BDF1 : C57BL$\times$DBA) and the DNA-injected eggs were treansferred into the oviducts of foster mothers. Genomic DNAs were isolated from the tails of mice born from the microinjected eggs and analyzed by Southern blot analysis. As a result, 5 and 9 transgenic mice with CChcLf and CChcLf-1 gene were produced, respectively. To determine tissue-specificity of transgene expression, Northern blot analysis was performed. Female transgenic mice were killed at day 10 of lactation and total RNAs from various tissues were isolated. Based on Northern blot analysis, it was shown that transgene was mainly expressed in the mammary glands of transgenic mice. In addition, the human lactoferrin in milk was detected by enzyme-linked immunosorbent assay. For this study, milk was obtained from the mammary glands of the transgenic mice at day 10 of lactation. In line #2 of CChcLf and line #7 of CChcLf-1 transgenic mice, human lactoferrin was secreted into the milk at concentration levels of 340ng/ml and 60ng/ml, respectively.

• Journal of Plant Biotechnology
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• 제7권1호
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• pp.45-50
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• 2005

To clone blue and white flower color specific genes, mRNA differential display was carried out with two different Commelina species, C. communis Linne for blue color and C. coreana Leveille for. leucantha Nakai for white color. Fifty two and 100 cDNA clones specific for blue or white flower color, respectively, were ranging from 200 to 700 bp in size. From the reverse northern blot analysis, 12 and 7 positive clones were selected for blue and white flower, respectively. These clones appear to be novel cDNAs for these Commelina plants, but not color specific. This finding was supported by the northern blot analysis. However, two clones, B18 and B19, derived from blue flowered Commelina were highly expressed than in the white Commelina species, implying that further study will be valuable. The results indicated that both mRNA display experiment and dot blot analysis may not sensitive enough to clone color-determining gene from the plant, leading to explore more advanced method, like high-density colony array study (HDCA).

• Journal of Plant Biotechnology
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• 제1권2호
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• pp.85-90
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• 1999

In order to protect fungal diseases, leaf disc explants of Solanum tuberosum cultivar, Belchip, was infected with an Agrobacterium MP90 strain containing chimeric gene construct, consisting of antibiotic resistance and chitinase gene driven by the CaMV 35S promoter, for transformation. Regenerated multiple shoots were selected on a medium containing kanamycin and carbenicillin after exposure to Agrobacterium. The presence and integration of the npt II and chitinase gene were confirmed by polymerase chain reaction(PCR). Northern blot analysis indicated that the genes coding for the enzyme could be expressed in potato plants. The chitinase activity of transgenic potato plants was higher than the control potato.

• 한국자원식물학회지
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• 제19권3호
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• pp.440-446
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• 2006

To investigate flowering related genes in winter-type oilseed rape (Brassica napus L. cv. Tammi), differentially expressed genes were isolated from leaves of the plant after low temperature treatment which is requirements for floral induction. As a result of suppression subtractive hybridization (SSH), 288 clones were randomly selected from SSH library. Using reverse Northern blot analysis, 150 of 288 clones were identified to be differentially expressed. Out of these 150 clones, 45 clones showed very high identities with the known genes. Four clones showed very high identities over 90% with metallothionein-like gene that is related to flowering-induced genes. Of these 4 clones, the cDNA clone, rfs-13, revealed high identity with meotallothionein-like protein in Arabidopsis thaliana (98%) and Brassica compestris (89%). Furthermore, gene expressed in immature flower stages was confirmed by Northern blot analysis.

• The Korean Journal of Physiology and Pharmacology
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• 제3권3호
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• pp.315-320
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• 1999

Molecular regulation of the renin-angiotensin system (RAS) was investigated in deoxycorticosterone acetate (DOCA)-salt hypertension. The expression of renin, angiotensinogen and angiotensin II receptor genes in the kidney and liver was determined by Northern blot analysis in rats which were made DOCA-salt hypertensive over the period of 2 or 4 weeks. Along with the hypertension, renin mRNA was decreased in the remnant kidney. The expression of angiotensinogen gene was not significantly altered in the kidney, but was significantly decreased in the liver. The expression of angiotensin II receptor gene was increased in the kidney, while it remained unaltered in the liver. The duration of hypertension did not affect the altered gene expression. It is suggested that the components of RAS are transcriptionally regulated in DOCA-salt hypertension in a tissue-specific manner.

• Journal of Plant Biotechnology
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• 제40권4호
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• pp.198-202
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• 2013

환경스트레스 저항성 오이 형질전환체 생산을 위해서 오이 "Eunsung" 품종의 자엽절 절편을 Nit유전자를 포함하는 pPZP211와 pCAMBIA2300 발현벡터로 각각 형질전환된 Agrobacterium과 공동 배양하였다. 공동배양 후 형질전환체 선발, 형질전환체 유도, 신장, 유식물체 생산 등은 자엽절 절편을 이용하는 CTM방법(Jang et al. 2011)에 따라 수행하였다. 발현벡터에 따라 선발배지에 100 mg/L paromomycin을 첨가하여 선발과정을 거쳤으며, 선발배지에서 3 cm크기의 shoot를 유도한 후 PCR, Southern, RT-PCR 및 Northern분석을 통해 형질전환 여부를 확인하였다. 공동배양 한 2,547개의 자엽절 절편으로부터 105개체(4.12%)가 선발배지로부터 얻어졌으며, 그들 중 45개체(1.77%)만이 Nit유전자의 PCR product를 얻을 수 있었다. 오이 genome에 Nit유전자의 도입여부를 확인하기 위하여 45개체에 대한 Southern분석을 수행한 결과 각각 39개체(1.53%)서 확인할 수 있었으며, 이중 오직 6개체(0.24%)에서만 Nit유전자가 안정적으로 발현되고 있음을 RT-PCR과 Northern분석을 통해 확인하였다. 이러한 결과는 Nit유전자가 오이 genome에 안정적으로 도입 및 발현되고 있음을 보여 주고 있음을 알 수 있었다.

• Plant Biotechnology Reports
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• 제3권4호
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• pp.277-283
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• 2009

One of the limitations to conducting maize Agrobacterium-mediated transformation using explants of immature zygotic embryos routinely is the availability of the explants. To produce immature embryos routinely and continuously requires a well-equipped greenhouse and laborious artificial pollination. To overcome this limitation, an Agrobacterium-mediated transformation system using explants of type II embryogenic calli was developed. Once the type II embryogenic calli are produced, they can be subcultured and/or proliferated conveniently. The objectives of this study were to demonstrate a stable Agrobacterium-mediated transformation of maize using explants of type II embryonic calli and to evaluate the efficiency of the protocol in order to develop herbicide-resistant maize. The type II embryogenic calli were inoculated with Agrobacterium tumefaciens strain C58C1 carrying binary vector pTF102, and then were subsequently cultured on the following media: co-cultivation medium for 1 day, delay medium for 7 days, selection medium for $4{\times}14$ days, regeneration medium, and finally on germination medium. The T-DNA of the vector carried two cassettes (Ubi promoter-EPSPs ORF-nos and 35S promoter-bar ORF-nos). The EPSPs conferred resistance to glyphosate and bar conferred resistance to phosphinothricin. The confirmation of stable transformation and the efficiency of transformation was based on the resistance to phosphinothricin indicated by the growth of putative transgenic calli on selection medium amended with $4mg\;1^{-1}$ phosphinothricin, northern blot analysis of bar gene, and leaf painting assay for detection of bar gene-based herbicide resistance. Northern blot analysis and leaf painting assay confirmed the expression of bar transgenes in the $R_1$ generation. The average transformation efficiency was 0.60%. Based on northern blot analysis and leaf painting assay, line 31 was selected as an elite line of maize resistant to herbicide.