共查询到20条相似文献,搜索用时 31 毫秒
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Cytoplasmic male sterility (CMS) in plants is known to be associated with novel open reading frames (ORFs) that result from
recombination events in the mitochondrial genome. In this study Southern and Northern blot analyses using several mitochondrial
DNA probes were conducted to detect the presence of differing band patterns between male fertile and CMS lines of chili pepper
(Capsicum annuum L.). In the CMS pepper, a novel ORF, termed orf456, was found at the 3′-end of the coxII gene. Western blot analysis revealed the expression of an approximately 17-kDa product in the CMS line, and the intensity
of expression of this protein was severely reduced in the restorer pepper line. To investigate the functional role of the
ORF456 protein in plant mitochondria, we carried out two independent experiments to transform Arabidopsis with a mitochondrion-targeted orf456 gene construct by Agrobacterium-mediated transformation. About 45% of the T1 transgenic population showed the male-sterile phenotype and no seed set. Pollen grains from semi-sterile T1 plants were observed to have defects on the exine layer and vacuolated pollen phenotypes. It is concluded that this newly
discovered orf456 may represent a strong candidate gene – from among the many CMS-associated mitochondrial genes – for determining the male-sterile
phenotype of CMS in chili pepper.
GenBank accession number DQ116040 (orf456 genomic sequence), DQ126683 (pepper coxII genomic sequence) 相似文献
5.
High-efficiency induction of soybean hairy roots and propagation of the soybean cyst nematode 总被引:8,自引:0,他引:8
Cotyledon explants of 10 soybean [Glycine max (L.) Merr.] cultivars were inoculated with Agrobacterium rhizogenes strain K599 with and without binary vectors pBI121 or pBINm-gfp5-ER possessing both neomycin phosphotransferase II (nptII) and β-glucuronidase (gus) or nptII and green fluorescent protein (gfp) genes, respectively. Hairy roots were produced from the wounded surface of 54–95% of the cotyledon explants on MXB selective
medium containing 200 μg ml−1 kanamycin and 500 μg ml−1 carbenicillin. Putative individual transformed hairy roots were identified by cucumopine analysis and were screened for transgene
incorporation using polymerase chain reaction. All of the roots tested were found to be co-transformed with T-DNA from the
Ri-plasmid and the transgene from the binary vectors. Southern blot analysis confirmed the presence of the 35S-gfp5 gene in the plant genomes. Transgene expression was also confirmed by histochemical GUS assay and Western blot analysis for
the GFP. Attempts to induce shoot formation from the hairy roots failed. Infection of hairy roots of the soybean cyst nematode
(Heterodera glycines Ichinohe)-susceptible cultivar, Williams 82, with eggs of H. glycines race 1, resulted in the development of mature cysts about 4–5 weeks after inoculation. Thus the soybean cyst nematode could
complete its entire life cycle in transformed soybean hairy-root cultures expressing GFP. This system should be ideal for
testing genes that might impart resistance to soybean cyst nematode.
Received: 13 July 1999 / Accepted: 8 August 1999 相似文献
6.
Ke Chen François Dorlhac de Borne Ernö Szegedi Léon Otten 《The Plant journal : for cell and molecular biology》2014,80(4):669-682
Nicotiana species carry cellular T‐DNA sequences (cT‐DNAs), acquired by Agrobacterium‐mediated transformation. We characterized the cT‐DNA sequences of the ancestral Nicotiana tabacum species Nicotiana tomentosiformis by deep sequencing. N. tomentosiformis contains four cT‐DNA inserts derived from different Agrobacterium strains. Each has an incomplete inverted‐repeat structure. TA is similar to part of the Agrobacterium rhizogenes 1724 mikimopine‐type T‐DNA, but has unusual orf14 and mis genes. TB carries a 1724 mikimopine‐type orf14‐mis fragment and a mannopine‐agropine synthesis region (mas2‐mas1‐ags). The mas2′ gene codes for an active enzyme. TC is similar to the left part of the A. rhizogenes A4 T‐DNA, but also carries octopine synthase‐like (ocl) and c‐like genes normally found in A. tumefaciens. TD shows a complex rearrangement of T‐DNA fragments similar to the right end of the A4 TL‐DNA, and including an orf14‐like gene and a gene with unknown function, orf511. The TA, TB, TC and TD insertion sites were identified by alignment with N. tabacum and Nicotiana sylvestris sequences. The divergence values for the TA, TB, TC and TD repeats provide an estimate for their relative introduction times. A large deletion has occurred in the central part of the N. tabacum cv. Basma/Xanthi TA region, and another deletion removed the complete TC region in N. tabacum. Nicotiana otophora lacks TA, TB and TD, but contains TC and another cT‐DNA, TE. This analysis, together with that of Nicotiana glauca and other Nicotiana species, indicates multiple sequential insertions of cT‐DNAs during the evolution of the genus Nicotiana. 相似文献
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Pawlicki-Jullian Nathalie Sedira Monika Welander Margareta 《Plant Cell, Tissue and Organ Culture》2002,70(2):163-171
The apple rootstock Jork 9 was transformed using four different Agrobacterium rhizogenes virulent strains. The mannopine strain 8196 gave the best results in the production of chimeric plants compared to two agropine strains (A4 and 15834) and one cucumopine strain. Shoot regeneration was performed on both untransformed and transformed roots. Optimum combination and concentration of thidiazuron (TDZ) and -naphtaleneacetic acid (NAA) was different between untransformed and transformed roots. From the transformed roots seven shoots were obtained and propagated as individual clones. All shoots from these clones rooted on a hormone-free medium contrary to untransformed shoots that did not root under similar culture conditions. Differences in the morphology of the leaves and stems were observed between the clones. The transformed status of the different clones was verified with mannopine tests, PCR and Southern blot analyses. Five clones contained the mas1', the ORF 13 and the rolB genes, whereas two clones contained only the rolB gene. 相似文献
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Factors that influence Agrobacterium rhizogenes-mediated transformation of broccoli (Brassica oleracea L. var. italica) 总被引:2,自引:0,他引:2
An improved broccoli transformation system was developed by optimising several factors that affect the rate of effective
Agrobacterium-mediated transformation. Leaf explants of cultivar Shogun were co-cultivated with Agrobacterium rhizogenes strain A4T harbouring the binary vector pART278. The T-DNA of this binary vector contains a neomycin phosphotransferase II
(NOS-NPTII-NOS) gene for kanamycin resistance and a β-glucuronidase (35S-GUS-OCS) gene. Several media and factors were evaluated
including combinations of arginine, mannopine, acetosyringone and the use of feeder cell layers. The new protocol includes
the use of 200 μm acetosyringone in LB medium for bacterial growth, the use of a Brassica campestris feeder cell layer, 10 mm mannopine and 50 μm acetosyringone in the co-cultivation medium and 1 mm arginine in the selection medium. The use of this optimised protocol produced transformation rates of 33% in preliminary
experiments transforming broccoli with the antisense 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase gene from pTOM13.
Received: 2 July 1998 / Revision received: 9 February 2000 / Accepted: 17 February 2000 相似文献
9.
Akutsu Masako Ishizaki Takuma Sato Hiroji 《Molecular breeding : new strategies in plant improvement》2004,13(1):69-78
An efficient procedure is described for transformation of calli of the monocotyledonous plant Alstroemeria by Agrobacterium rhizogenes. Calli were co-cultivated with A. rhizogenes strain A13 that harbored both a wild-type Ri-plasmid and the binary vector plasmid pIG121Hm, which included a gene for neomycin phosphotransferase II (NPTII) under the control of the nopaline synthase (NOS) promoter, a gene for hygromycin phosphotransferase (HPT) under the control of the cauliflower mosaic virus (CaMV) 35S promoter, and a gene for -glucuronidase (GUS) with an intron fused to the CaMV 35S promoter. Inoculated calli were plated on medium that contained cefotaxime to eliminate bacteria. Four weeks later, transformed cells were selected on medium that contained 20 mg L–1 hygromycin. A histochemical assay for GUS activity revealed that selection by hygromycin was complete after eight weeks. The integration of the T-DNA of the Ri-plasmid and pIG121Hm into the plant genome was confirmed by PCR. Plants derived from transformed calli were produced on half-strength MS medium supplemented with 0.1 mg L–1 GA3 after about 5 months of culture. The presence of the gusA, nptII, and rol genes in the genomic DNA of regenerated plants was detected by PCR and Southern hybridization, and the expression of these transgenes was verified by RT-PCR. 相似文献
10.
The wheat mitochondrial orf25 nucleotide sequence of 576 pb has been determined. Its derived protein sequence shares 88% and 75% amino acid identity with those of maize and tobacco mitochondria, respectively. The wheat and tobacco orf25 sequences lack four inserts, of 6 bp to 36 bp, that are present in the maize homologue. The wheat orf25 gene is actively transcribed and is preceded by a regulatory sequence block very similar to those located upstream of the wheat coxII and atp6 genes. Our observations support the view that orf25 sequences encode a functional polypeptide in plant mitochondria. 相似文献
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SS Swain L Sahu A Pal DP Barik C Pradhan PK Chand 《World journal of microbiology & biotechnology》2012,28(2):729-739
Transformed rhizoclones were developed from Agrobacterium-treated explants of the medicinally important twinning legume Clitoria ternatea L. Several key factors influencing transformation events were optimized. A4T was the most infectious among the strains employed.
Internode segments were more responsive than leaves, outdoor-grown explants preferred to those from in vitro cultures. High
frequency transformation, resulting in up to 85.8% rhizogenesis, was attained using pre-pricked internodal explants for immersion
(10 min) in Agrobacterium rhizogenes suspension grown overnight with acetosyringone (100 μM) to an OD660 ≅ 0.6, diluted to a density of 109 cells ml−1, followed by 5-day co-cultivation. Roots were individually cultured in MS0 supplemented with the bacteriostatic antibiotic
cefotaxime (500 μg ml−1). Rhizoclones were renewed through successive subcultures in MS0 under diffused illumination. The T
L
-DNA rolB and rolC ORF were detected in rhizoclones through PCR amplification. The T
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-DNA gene encoding mannopine synthase (man2) was revealed by positive amplification and opine gene expression substantiated by agropine and mannopine biosynthesis in
all selected transformed rhizoclones. The implication of such findings is discussed on the context of utilization of such
genetically transformed root cultures towards sustainable production of medicinally useful phytocompounds, besides providing
a means for plant conservation. 相似文献
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Tissue-specific expression of the ORF13 promoter from Agrobacterium rhizogenes 8196 was assessed throughout the development of transgenic tobacco plants using a GUS reporter gene. ORF13 exhibited high activity in roots but with different patterns of expression. The activity of the ORF13 promoter in vascular tissues increased from the base to the tip of the stem. The ORF13 promoter is wound inducible in a limited area adjacent to the wound site. The time course of wound induction of ORF13 in transgenic tobacco containing an ORF13 promoter-GUS translational fusion was similar to that previously described for genes involved in plant defense responses. A series of 5′ deletions of the ORF13 promoter fused to the β-glucuronidase gene was examined for expression in roots and leaves of transgenic plants. Cis-acting elements that modulate quantitative expression of the transgene after wounding were detected. 相似文献
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Tissue-specific expression of the ORF13 promoter from Agrobacterium rhizogenes 8196 was assessed throughout the development of transgenic tobacco plants using a GUS reporter gene. ORF13 exhibited high
activity in roots but with different patterns of expression. The activity of the ORF13 promoter in vascular tissues increased
from the base to the tip of the stem. The ORF13 promoter is wound inducible in a limited area adjacent to the wound site.
The time course of wound induction of ORF13 in transgenic tobacco containing an ORF13 promoter-GUS translational fusion was
similar to that previously described for genes involved in plant defense responses. A series of 5′ deletions of the ORF13
promoter fused to the β-glucuronidase gene was examined for expression in roots and leaves of transgenic plants. Cis-acting
elements that modulate quantitative expression of the transgene after wounding were detected.
Received: 11 July 1996 / Accepted: 19 November 1996 相似文献
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Seishirō Aoki Akiyoshi Kawaoka Masami Sekine Takanari Ichikawa Tomomichi Fujita Atsuhiko Shinmyō Kunihiko Syōno 《Molecular & general genetics : MGG》1994,243(6):706-710
A region homologous to the TL-DNA of Agrobacterium rhizogenes was previously detected in the genome of untransformed Nicotiana glauca and designated cellular T-DNA (cT-DNA). Subsequently, part of this region was sequenced and two genes, which corresponded to rolB and rolC and were named NgrolB and NgrolC, were found. We have now sequenced a region of the cT-DNA other than the region that includes NgrolB and C and we have found two other open reading frames (ORFs), NgORF13 and NgORF14. These ORFs correspond to ORFs 13 and 14 of the TL-DNA of A. rhizogenes and exhibit a high degree of homology to these ORFs, without having a nonsense codon. We have not found any sequence homologous to rolD (ORF15). The two genes, NgORF13 and 14, as well as the NgrolB and C genes, are expressed in genetic tumors of hybrids between N. glauca and N. langsdorffii but not in leaf tissues of the hybrid. 相似文献
15.
Acinetobacter calcoaceticus PHEA-2 is a phenol-degrading bacterium isolated from the wastewater from an oil refinery. A 10-kb XhoI fragment consisting of nine complete Open Reading Frames (ORFs) and one partial ORF was screened from a lambda library of
PHEA-2 by Southern hybridization. The sequence analyses revealed that ORF2–ORF7, designated mphKLMNOP, are homologous to dmpKLMNOP of Pseudomonas sp. CF600 and mopKLMNOP of Acinetobacter calcoaceticus NCIB8250, sharing 38%–72% and 58.5%–93.5% respectively. The products encoded by dmp and mop genes convert phenol to catechol. The mph-operon and downstream ORFs, ORF9 and ORF10, sharing high identities to benM and benA, which encode ben-operon regulatory protein and benzoate 1,2-dioxygenase alpha subunit respectively, are separated by ORF8, whose function
is unknown. The organization of the mph and ben operons is different from that described previously.
Received: 8 April 2002 / Accepted: 8 May 2002 相似文献
16.
José Luis Cabrera-Ponce Ariadne Vegas-Garcia Luis Herrera-Estrella 《In vitro cellular & developmental biology. Plant》1996,32(2):86-90
AnAgrobacterium rhizogenes-mediated procedure for transformation of papaya (Carica papaya) was developed. Transgenic plants were obtained from somatic embryos that spontaneously formed at the base of transformed
roots, induced from leaf discs infected withA. rhizogenes. Transformation was monitored by autonomous growth of roots and somatic embryos, resistance to kanamycin, β-glucuronidase
activity (GUS), and Southern hybridization analysis. Over one-third of the infected leaf explants produced transformed roots
with GUS activity, from which 10% spontaneously produced somatic embryos. Histological analysis ofA. rhizogenes-transformed embryos showed that they have an altered symmetry between the shoot apex and the root meristem when compared
to somatic embryos induced with hormone treatment from control explants. Transgenic papaya plants containingA. rhizogenes rol genes were more sensitive to auxins, developed wrinkled leaves, and grew slower than nontransformed plants. 相似文献
17.
E. A. Trifonova A. V. Romanova S. S. Sangaev M. V. Sapotsky V. I. Malinovsky A. V. Kochetov 《Biologia Plantarum》2012,56(3):571-574
The gene of Zinnia elegans L. coding for S-like extracellular ribonuclease (ZRNase II) was used to produce transgenic tobacco plants with an increased ribonuclease activity. The protein-coding part of ZRNase
II included the signal peptide sequence so the transgenic protein was located extracellularly. The cDNA of ZRNase II was cloned under the control of 2′-promoter of the mannopine synthase (MAS 2′) gene from Ti-plasmid of Agrobacterium tumefaciens. It was shown that the resultant transgenic plants had an increased ribonuclease activity of the crude extracts and the induction
of MAS 2′ promoter by wounding additionally increased the activity. The plants of two transforming lines characterized by
different ribonuclease activities were used to analyze the transgene influence on plant resistance to tobacco mosaic virus.
The plants demonstrated either absence of disease symptoms or a significant delay in their appearance, depending on the virus
content in the inoculum and ribonuclease activity. 相似文献
18.
The RT PCR and RACE methods were used to obtain the cDNA sequence of an APX gene of muskmelon after the leaves were induced
with powdery mildew. The cDNA length of the APX gene is 1,047 bp with a 750 bp ORF encoded a 249 amino acid and the molecular
weight of APX protein is 27.3 kDa. The analysis showed that the CmAPX genomic DNA contained 10 extrons and 9 introns. The identity of the amino acid sequence deduced from the cDNA with the
APX family of other homologous members was about 74–97%. A Full-length of ORF was sub-cloned into prokaryotic expression vector
pET24a. The recombinant proteins had high expression level in E. coli. Analysis of expression at mRNA level showed that CmAPX exhibited highly tissue-specific patterns of expression. The mRNA level and enzyme activities assays showed that CmAPX might play an important role in the pathogenesis of powdery mildew. 相似文献
19.
E. Boudreau M. Turmel M. Goldschmidt-Clermont J.-D. Rochaix S. Sivan A. Michaels S. Leu 《Molecular & general genetics : MGG》1997,253(5):649-653
An open reading frame potentially encoding a protein of 1995 amino acids (orf1995) has been found in the chloroplast genome of the green alga Chlamydomonas reinhardtii. Besides having a short hydrophobic N-terminal domain with five putative transmembrane helices, the predicted orf1995 product is highly basic. orf1995 might be a homologue of the ycf1 gene in land plants, whose function has not yet been determined. Mutants of C. reinhardtii transformed with a disruption of orf1995 remain heteroplasmic for the wild-type and disrupted alleles of this gene, indicating that the orf1995 product is essential for cell survival.
Received: 18 August 1996 / Accepted: 24 September 1996 相似文献