• Journal of Plant Biology
• /
• 제33권2호
• /
• pp.97-104
• /
• 1990

In order to investigate the host range of Agrobacterium tumefaciens KU12 containing pTi-12, 28 species of dicotyledonous plants were infected with KU12, A136 without Ti plasmid and A348 containing pTi A6, respectively. KU12 and A348 induced tumor in 20 species and 14 species, respectively. This results showed that KU12 has a wide host range. Therefore, it was confirmed that KU12 and pTi-12 are very useful for developing plant vector system having a broad host range.

• 한국미생물·생명공학회지
• /
• 제18권1호
• /
• pp.94-97
• /
• 1990

Thiostrepton 내성 유전자(tsr)를 포함하는 multicopy 재조합 플라스미드 pJY502(5.5kb)의 제한효소 지도를 비교해본 결과 pJY502는 새로운 플라스미드로 확인되었다. pJY502는 Streptomyces에서 넓은 host range를 나타내었으며 cloning에 사용할 수 있는 단일 BglII 제한효소 인식부위를 갖고 있었다. pJY502의 형질전환 빈도는 S.lividans에서 $2.2 \times 10^5$이었다. 또한 E.coli-Streptomyces bifunctional 플라스미드 pJY504을 제조하였다.

• Journal of Microbiology and Biotechnology
• /
• 제1권4호
• /
• pp.240-245
• /
• 1991

We have constructed a promoter-probe vector pKU20 using pKT230, a derivative of broad-host-range plsmid RSF1010, as a base. The pKU20 contains structural gene for aminoglycoside phos-photransferase (aph), without promoter, and a multiple cloning site upstream the aph. Using this vector, a 412base pairs (bp) PstI fragment showing strong promoter activity both in Escherichia coli LE392 and Pseudomonas putida KCTC1644 has been cloned from Pseudomonas fluorescens chromosomal DNA on the basis of streptomycin resistance. The nucleotide sequence of the 412 bp fragment has been determined and the putative - 35 and -10 region was observed. Insecticidal protein gene of Bacillus thuringiensis subsp. kurstaki HD-73 inserted on downstream of the promoterlike DNA fragment was efficiently expressed in E. coli and P. putida. The toxin protein was efficiently synthesized in an insoluble form in both strains.

• 미생물학회지
• /
• 제28권1호
• /
• pp.35-40
• /
• 1990

Thiostrepton 내성 유전자(tsr)를 포함하는 multi-copy 재조합 플라스미드 pJY7J2의 제한효소 절단지도를 작성하였다. pJY, 712는 Streptomyces에서 넓은 host range를 나타내었으며 cloning 목적에 사용할 수 있는 단일 BgtIl 제한효소 인식부위를 갖고 있었다. 플라스미드 pJY 712는 lethal zygosis(Ltz+) 현상을 보였다. pJY 712의 혁질전환빈도는 S. lividans에서 $5.0\times 10^{4}$ TFU였다. pJY 712의 Bell 제한효소 인식부위에 tyrosmase 유전자(mel)를 삽입하여 플라스미드 PJY713을 제조하였다. met 유전자를 포함한 재조합 플라스미드 pJY 714는 pJY 713의 일부분(1.9kb BgllI-BelI 단편)을 제거하여 제고하였다.

• 미생물학회지
• /
• 제22권4호
• /
• pp.249-255
• /
• 1984

To develop the host-vector system for industrial Coryneform bacteria that seemed to be the most suitable microorganisms for molecular breeding of genes involved in the production of amion acids, nucleotides, and other products of industrial interest, broad host range E. coli plasmid R 1162 DNA was transformed into Brevibacterium ammoniagenes and the plasmids pKU6 isolated from a transformant was physically characterized. All other plasmids from the transformed cells except pKU6 exsisted as multimeric forms in Brevibacterium ammoniagenes. The plasmid DNA was retransformed into Corynebacterium glutamicum with a high frequency ($1.32{\times}10^{-1}$ per cell) and maintained stably both in Brevibacterium ammoniagenes and Corynebacterium glutamicum after 100 generations of cultures with 25-30 copy number per cell. The size of both plasmid pKU6 and plasmid R1162 were the same and restriction maps by EcoR I, Ava I, Pst I, Pvu II and Hinc II were also similar.

• 미생물학회지
• /
• 제27권2호
• /
• pp.92-97
• /
• 1989

the vector, pGUR19, for Rhizobium gene manipulation, was constructed by combining the replication and mobilization function of indigenous multicopy plasmid from Acacia(Robinia pseudoacacia L.) Rhizobia sp86 with E. coli cloning vehicle, pBR322. The vector could be efficiently mobilized by RP4 tra function incorporated into chromosome of E. coli named SM10 and efficiently transferred to various gram negative hosts including Rhizobium and Afrobacterium by transformation. Mobilization frequency of the constructed vector was ranged from $1.2\times 10^{-2}$ (E.coli HB 101) to $4.6\times 10^{-4}$ (A. tumefaciens 15955) and transformation frequency was ranged from $5.4\times 10^{-7}$(E. coli HB101) to $1.2\times 10^{-10}$ (A. tumefaciens 15955). The vector, pGUR19, was stably replicated and maintained in a variety of Rhizobium and Agrobacterium.

• Journal of Microbiology and Biotechnology
• /
• 제21권11호
• /
• pp.1123-1126
• /
• 2011

The IncP plasmid R995 has been a useful self-transmissible, broad-host-range vector for a number of applications including the recombinase/conjugation-based cloning of large genomic DNA segments. However, R995 derivatives (or related plasmids) expressing a wide range of different resistance markers and Flp recombinase target sites do not exist in the literature. In addition, documented strategies for applying such plasmids in cloning applications that take advantage of conjugation for the convenient isolation and recovery of constructs are extremely limited. Here, we report a new series of R995 plasmids with alternative markers to increase options for applications in backgrounds already expressing resistance to a particular antibiotic(s). These R995 plasmids have been engineered to contain FRT sites that can be used for recombinase-based cloning. We demonstrate the utility of this approach by cloning 20 kb regions from the Salmonella Typhimurium and Escherichia coli genomes and by cloning DNA from an exogenous plasmid source. To our knowledge, this represents the first systematic engineering of an intact, self-transmissible IncP plasmid with a series of alternative antibiotic markers and FRT sites.

• KSBB Journal
• /
• 제11권1호
• /
• pp.115-123
• /
• 1996

zooglan생합성에 필수적인 gene cluster를 clone하기 위해 2종류의 변이주가 분리되었다. Zoogloea ramigera 115는 협막형 다당(주로 zooglan)을 생산한다. 115 균주로 접합시키고 생산물을 용이하게 분리하기 위하여 반복된 원심분리와 선별을 통해 협막을 만들지 않는 slime형 생산균주를 분리하였다. 세포외 다당 생산 능력이 결여된 변이주를 전통적인 transposon(Tn5) 기술을 사용하여 얻었고 달라진 colony 형태와 celluflour결합 성질에 의해 선별하였다. 이들 변이주들은 범용숙주범위 cosmid vector안에 건설된 Z.ramigera 115slime gene library 와 helper plasmid로의 3양친 접합에 의해 보상되었다.

• 농업과학연구
• /
• 제44권2호
• /
• pp.145-180
• /
• 2017

Alternanthera mosaic virus (AltMV) is a member of the genus Potexvirus which has been known for less than twenty years, and has been detected in Australasia, Europe, North and South America, and Asia. The natural host range to date includes species in at least twenty-four taxonomically diverse plant families, with species in at least four other families known to be infected experimentally. AltMV has been shown to differ from Potato virus X (PVX), the type member of the genus Potexvirus, in a number of ways, including the subcellular localization of the Triple Gene Block 3 (TGB3) protein and apparent absence of interactions between TGB3 and TGB2. Differences between AltMV variants have allowed identification of viral determinants of pathogenicity, and identification of residues involved in interactions with host proteins. Infectious clones of AltMV differing significantly in symptom severity and efficiency of RNA silencing suppression have been produced, suitable either for high level protein expression (with efficient RNA silencing suppression) or for Virus-Induced Gene Silencing (VIGS; with weaker RNA silencing suppression), demonstrating a range of utility not available with most other plant viral vectors. The difference in silencing suppression efficiency was shown to be due to a single amino acid residue substitution in TGB1, and to differences in subcellular localization of TGB1 to the nucleus and nucleolus. The current state of knowledge of AltMV biology, including host range, strain differentiation, host interactions, and utility as a plant viral vector for both protein expression and VIGS are summarized.

• 대한수의학회지
• /
• 제29권3호
• /
• pp.407-416
• /
• 1989

The prospect of live vaccines consisting of genetically modified vaccinia virus expressing foreign genes is exciting, but important issues concerning safety and efficacy need to resolved. Vaccinia virus (VV) is an efficient expression vector with broad host range infectivity and large DNA capacity. This vector has been particularly useful for identifying target antigens for humoral and cell-mediated immunity. The WHO smallpox eradication program, involving the extensive use of VV vaccines, resulted in the late 1970s in the elimination of one of the world's most feared diseases. This achievement is a triumph for preventive medicine and for international collaboration in public health. In 1980, WHO recommended that the routine use of smallpox vaccine should be stopped. Against this background, the prospect of li ve vaccines consisting of genetically modified VV expressing foreign antigens arising from the work of Moss, and Paoletti and their colleagues in 1982 has been greeted with enthusiasm. These investigators have shown that genes coding for immunogenic proteins can be inserted into VV DNA without impairing the ability of the virus to grow in cell culture. Moreover experimental animals infected with VV recombinants containing genes coding for a variety of immunizing proteins have been shown to be protected against challenge infection with the corresponding infectious agent. In this communication, I describe current progress in the construction of a novel plasmid vector that facilitate the insertion and expression of foreign genes in VV as well as the selection of recombinants.