植物学技术

The Potyviridae are the largest single group of plant viruses, and as such are the most important from an economic standpoint. There are nearly 200 distinct recognized species or other viruses that are possible or probable members of the group. Two recent books provide an excellent and in depth rev ...

All members of the genus are known to be transmitted mechanically, with the majority also being transmitted in a nonpersistent manner by aphids (1), though one confirmed carlavirus is known to be transmitted by whiteflies (2). Carlaviruses are noted for their narrow host range and tendency to in ...

The potexvirus family has at least 30 definitive and possible members, of which potato virus X (PVX) is the type member (1–4). All potexviruses are morphologically similar, with flexuous, filamentous virions that range from 470 to 580 nm in length. Each virus particle contains a single-stranded, ...

In the first three decades of the 20th century, it was shown that a number of plant diseases could be transmitted by infectious sap that had been passed through a bacteria-proof filter. Plant virus particles had yet to be identified and characterized, and much of the research effort of plant virologis ...

The tobamoviruses produce rigid, rod-shaped particles, with dimensions of approx 300 � 18 nm, and form one of the most extensively studied groups of plant viruses (1). Members of the group infect a wide range of angiosperms, and individual members frequently have wide experimental host ranges. T ...

The tobraviruses have a genome consisting of two positive-sense, single-stranded RNA molecules. The two genomic RNAs, RNA-1 and RNA-2, are encapsidated separately in rod-shaped particles with lengths of 180–215 nm (L particles) and 46–115 nm (S particles), respectively (Fig. 1) (1,2). Both the ...

The Furovirus (fungus-transmitted rod-shaped) group, whose type member is soil-borne wheat mosaic virus (SBWMV) (1), is a very heterogeneous group infecting a wide variety of hosts. The group consists of viruses that are naturally transmitted by soil-borne fungal vectors, generally of t ...

Agrobacterium-mediated transfer of cloned DNA copies of infectious viral genomes to plant tissues has been termed “agroinoculation” or “agroinfection” (for review, see ref. 1). The technique has been used successfully for a wide variety of virus groups, including the caulimoviruses a ...

Introduction of foreign genes into plant cells can be achieved by a variety of methods, including direct transfer into protoplasts using chemical (1–3; see also Chapter 7, this volume) and electrical methods (4–6). These methods can be used to study genes both transiently (1, 5,7) and when stably int ...

The cereals are one of the groups of crops more recalcitrant to transformation. Since, with the exception of rice (1), cereals have not been transformed by Agrobacterium, and highly regenerable protoplast systems are difficult to obtain, these species have remained untransformed for a much ...

The chloramphenicol acetyltransferase (cat) gene, isolated from transposon Tn9 of Escherichia coli (1), is a convenient genetic marker for studies of transformation. (For a detailed description of the CAT gene, see Chapter 1.) In plants, the CAT system is generally used in transient, rather t ...

The stable introduction of genes into plants through genetic engineering normally necessitates the use of a selectable marker, especially when the transformation frequency is low (e.g., 1.0 � 10−3 to 10−6). Marker genes enable quantification of both the transformation efficiency and ge ...

The study of how genes are organized within the genome is a major step in the characterization of cloned DNA sequences and is important in the analysis of plant transformation. In both cases, accurate information is required about the number of copies of a given DNA sequence and their location in the gen ...

Today the isolation and characterization of a gene of interest from any organism has become a standard procedure. One of the milestones that have facilitated this procedure in particular, has been the development of in vitro techniques for amplification of specific RNA or DNA sequences. By far ...

Eukaryotic messenger RNA (mRNA) can be separated from the other RNA species in a total RNA preparation by affinity chromatography by virtue of the presence of a polyadenylic acid “tail,” 20–25 bases long, at the 3′ end of the molecule (1). Oligo(dT)-cellulose is used routinely for homemade affinity c ...

Efficient extraction of high quality RNA from a variety of plant tissues is an important first step in many procedures, such as analysis of gene expression, cDNA library construction, and in vitro translation. The procedure described here, which is essentially the same as described in Draper et ...

Soon after the introduction of the Southern blotting procedure (1), an analogous technique, known as northern blotting, was developed for the analysis of RNA sequences (2). Both procedures rely on the annealing (hybridization) of complementary single-stranded nucleic acid molecul ...

Nuclear “run-on” (or “run-off”) transcription assays have been used to obtain quantitative information about the relative rates of transcription of different genes in nuclei isolated from a particular tissue or organ. This information can then be used in analyzing the factors that contr ...

RNase A/T1 mapping provides a sensitive and quantitative method of expression analysis of known gene sequences. It differs from primer-derived analyses such as primer extension and reverse transcriptase-PCR by the use of a probe that is colinear with the transcript under study. This prot ...

A number of protocols are available for the analysis of RNA transcripts: Northern analysis (see Chapter 18), S1 mapping, primer extension, RNase AT1 mapping (see Chapter 20), and the more recent reverse transcriptase/PCR amplification (see Chapters 22 and24). These protocols have been app ...