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1.  Spatial and temporal transcriptome changes occurring during flower opening and senescence of the ephemeral hibiscus flower, Hibiscus rosa-sinensis  
Journal of Experimental Botany  2016;67(20):5919-5931.
Pathway analysis suggests that identified temporal and spatial transcriptomic changes associated with senescence of the short-lived hibiscus flower are regulated by light/circadian clock-, aquaporin-, cell wall-, and calcium-related gene families.
Flowers are complex systems whose vegetative and sexual structures initiate and die in a synchronous manner. The rapidity of this process varies widely in flowers, with some lasting for months while others such as Hibiscus rosa-sinensis survive for only a day. The genetic regulation underlying these differences is unclear. To identify key genes and pathways that coordinate floral organ senescence of ephemeral flowers, we identified transcripts in H. rosa-sinensis floral organs by 454 sequencing. During development, 2053 transcripts increased and 2135 decreased significantly in abundance. The senescence of the flower was associated with increased abundance of many hydrolytic genes, including aspartic and cysteine proteases, vacuolar processing enzymes, and nucleases. Pathway analysis suggested that transcripts altering significantly in abundance were enriched in functions related to cell wall-, aquaporin-, light/circadian clock-, autophagy-, and calcium-related genes. Finding enrichment in light/circadian clock-related genes fits well with the observation that hibiscus floral development is highly synchronized with light and the hypothesis that ageing/senescence of the flower is orchestrated by a molecular clock. Further study of these genes will provide novel insight into how the molecular clock is able to regulate the timing of programmed cell death in tissues.
PMCID: PMC5091337  PMID: 27591432
454 sequencing; aquaporins; cell wall; light and circadian clock signalling; microarray; ovary; PCD; petal; style-stigma plus stamens.
2.  A Narcissus mosaic viral vector system for protein expression and flavonoid production 
Plant Methods  2013;9:28.
With the explosive numbers of sequences generated by next generation sequencing, the demand for high throughput screening to understand gene function has grown. Plant viral vectors have been widely used as tools in down-regulating plant gene expression. However, plant viral vectors can also express proteins in a very efficient manner and, therefore, can also serve as a valuable tool for characterizing proteins and their functions in metabolic pathways in planta.
In this study, we have developed a Gateway®-based high throughput viral vector cloning system from Narcissus Mosaic Virus (NMV). Using the reporter genes of GFP and GUS, and the plant genes PAP1 (an R2R3 MYB which activates the anthocyanin pathway) and selenium-binding protein 1 (SeBP), we show that NMV vectors and the model plant Nicotiana benthamiana can be used for efficient protein expression, protein subcellular localization and secondary metabolite production.
Our results suggest that not only can the plant viral vector system be employed for protein work but also can potentially be amenable to producing valuable secondary metabolites on a large scale, as the system does not require plant regeneration from seed or calli, which are stages where certain secondary metabolites can interfere with development.
PMCID: PMC3728148  PMID: 23849589
3.  Colour break in reverse bicolour daffodils is associated with the presence of Narcissus mosaic virus 
Virology Journal  2011;8:412.
Daffodils (Narcissus pseudonarcissus) are one of the world's most popular ornamentals. They also provide a scientific model for studying the carotenoid pigments responsible for their yellow and orange flower colours. In reverse bicolour daffodils, the yellow flower trumpet fades to white with age. The flowers of this type of daffodil are particularly prone to colour break whereby, upon opening, the yellow colour of the perianth is observed to be 'broken' into patches of white. This colour break symptom is characteristic of potyviral infections in other ornamentals such as tulips whose colour break is due to alterations in the presence of anthocyanins. However, reverse bicolour flowers displaying colour break show no other virus-like symptoms such as leaf mottling or plant stunting, leading some to argue that the carotenoid-based colour breaking in reverse bicolour flowers may not be caused by virus infection.
Although potyviruses have been reported to cause colour break in other flower species, enzyme-linked-immunoassays with an antibody specific to the potyviral family showed that potyviruses were not responsible for the occurrence of colour break in reverse bicolour daffodils. Colour break in this type of daffodil was clearly associated with the presence of large quantities of rod-shaped viral particles of lengths 502-580 nm in tepals. Sap from flowers displaying colour break caused red necrotic lesions on Gomphrena globosa, suggesting the presence of potexvirus. Red necrotic lesions were not observed in this indicator plant when sap from reverse bicolour flowers not showing colour break was used. The reverse transcriptase polymerase reactions using degenerate primers to carla-, potex- and poty-viruses linked viral RNA with colour break and sequencing of the amplified products indicated that the potexvirus Narcissisus mosaic virus was the predominant virus associated with the occurrence of the colour break.
High viral counts were associated with the reverse bicolour daffodil flowers that were displaying colour break but otherwise showed no other symptoms of infection. Narcissus mosaic virus was the virus that was clearly linked to the carotenoid-based colour break.
PMCID: PMC3170305  PMID: 21854646
4.  Methods for transient assay of gene function in floral tissues 
Plant Methods  2007;3:1.
There is considerable interest in rapid assays or screening systems for assigning gene function. However, analysis of gene function in the flowers of some species is restricted due to the difficulty of producing stably transformed transgenic plants. As a result, experimental approaches based on transient gene expression assays are frequently used. Biolistics has long been used for transient over-expression of genes of interest, but has not been exploited for gene silencing studies. Agrobacterium-infiltration has also been used, but the focus primarily has been on the transient transformation of leaf tissue.
Two constructs, one expressing an inverted repeat of the Antirrhinum majus (Antirrhinum) chalcone synthase gene (CHS) and the other an inverted repeat of the Antirrhinum transcription factor gene Rosea1, were shown to effectively induce CHS and Rosea1 gene silencing, respectively, when introduced biolistically into petal tissue of Antirrhinum flowers developing in vitro. A high-throughput vector expressing the Antirrhinum CHS gene attached to an inverted repeat of the nos terminator was also shown to be effective. Silencing spread systemically to create large zones of petal tissue lacking pigmentation, with transmission of the silenced state spreading both laterally within the affected epidermal cell layer and into lower cell layers, including the epidermis of the other petal surface. Transient Agrobacterium-mediated transformation of petal tissue of tobacco and petunia flowers in situ or detached was also achieved, using expression of the reporter genes GUS and GFP to visualise transgene expression.
We demonstrate the feasibility of using biolistics-based transient RNAi, and transient transformation of petal tissue via Agrobacterium infiltration to study gene function in petals. We have also produced a vector for high throughput gene silencing studies, incorporating the option of using T-A cloning to insert the gene sequence of interest. These techniques should allow analysis of gene function in a much broader range of flower species.
PMCID: PMC1781449  PMID: 17207290
7.  Isolation of Bacteriophages Active Against Neisseria meningitidis 
Journal of Virology  1967;1(3):538-542.
Five distinct bacteriophages have been isolated from strains of Neisseria meningitidis. Filtrates with titers of 10−4 to 10−6 were produced with a modified Swanstrom and Adams semisolid agar procedure, employing Eugonbroth with added agar and an incubation temperature of 30 C. Of 49 strains of N. meningitidis (groups B and C), 25 were lysed by one or more of the phages, but there was no lysis of other Neisseria and Mima polymorpha strains.
PMCID: PMC375274  PMID: 4990042
9.  Harveian Oration 
British Medical Journal  1957;2(5052):1047-1048.
PMCID: PMC1962691
11.  Sayings of Robert Hutchison 
Archives of Disease in Childhood  1951;26(129):467-468.
PMCID: PMC1988430  PMID: 14886057
12.  Mercurialentis * 
PMCID: PMC1324121  PMID: 13032384
17.  Protection of the Worker Against Injury and Disease.—II* 
British Medical Journal  1950;1(4652):506-512.
PMCID: PMC2037289  PMID: 20787763
19.  Protection of the Worker Against Injury and Disease.—I* 
British Medical Journal  1950;1(4651):449-454.
PMCID: PMC2036973  PMID: 15405906
21.  Occupational Medicine 
British Medical Journal  1949;1(4595):184.
PMCID: PMC2049452

Results 1-25 (43)