Additional file 1 of Recombinant-attenuated Salmonella enterica serovar Choleraesuis vector expressing the PlpE protein of Pasteurella multocida protects mice from lethal challenge

Supplementary Material

Additional file 2 of Recombinant-attenuated Salmonella enterica serovar Choleraesuis vector expressing the PlpE protein of Pasteurella multocida protects mice from lethal challenge

Supplementary Material

Upland Cotton Gene <i>GhFPF1</i> Confers Promotion of Flowering Time and Shade-Avoidance Responses in <i>Arabidopsis thaliana</i>

<div><p>Extensive studies on floral transition in model species have revealed a network of regulatory interactions between proteins that transduce and integrate developmental and environmental signals to promote or inhibit the transition to flowering. Previous studies indicated <i>FLOWERING PROMOTING FACTOR 1</i> (<i>FPF1</i>) gene was involved in the promotion of flowering, but the molecular mechanism was still unclear. Here, <i>FPF1</i> homologous sequences were screened from diploid <i>Gossypium raimondii</i> L. (D-genome, n = 13) and <i>Gossypium arboreum</i> L. genome (A-genome, n = 13) databases. Orthologous genes from the two species were compared, suggesting that distinctions at nucleic acid and amino acid levels were not equivalent because of codon degeneracy. Six <i>FPF1</i> homologous genes were identified from the cultivated allotetraploid <i>Gossypium hirsutum</i> L. (AD-genome, n = 26). Analysis of relative transcripts of the six genes in different tissues revealed that this gene family displayed strong tissue-specific expression. <i>GhFPF1</i>, encoding a 12.0-kDa protein (Accession No: KC832319) exerted more transcripts in floral apices of short-season cotton, hinting that it could be involved in floral regulation. Significantly activated <i>APETALA 1</i> and suppressed <i>FLOWERING LOCUS C</i> expression were induced by over-expression of <i>GhFPF1</i> in the <i>Arabidopsis</i> Columbia-0 ecotype. In addition, transgenic <i>Arabidopsis</i> displayed a constitutive shade-avoiding phenotype that is characterized by long hypocotyls and petioles, reduced chlorophyll content, and early flowering. We propose that <i>GhFPF1</i> may be involved in flowering time control and shade-avoidance responses.</p></div

Gene structure of <i>GhFPF1</i>, and its response to plant hormones.

<p>A. The gene structure of <i>GhFPF1</i>. JA and SA represent response elements of JA and SA; TSS, transcriptional start site. B. <i>GhFPF1</i> and <i>GhNPR1</i> expression profiles in the first forty-eight hours after treatment with SA and JA. <i>GhNPR1</i> (DQ409173), a known JA- and SA-inducible gene, was used as a positive control here. Error bars represent SD.</p

Expression patterns of <i>GhFPF1</i>, <i>GhFLP-1</i>, <i>GhFLP-2</i>, <i>GhFLP-3</i>, <i>GhFLP-4</i>, and <i>GhFLP-5</i> in <i>G. hirsutum</i> L.

<p>Relative expression of <i>GhFPF1</i> and its homologs were measured in different tissues of upland cotton CCRI 36 (a short-season cotton variety) and TM-1 (a genetic standard line) using qRT-PCR. Roots, stems, leaves, flowers and fibers stand for mixed samples from CCRI 36 and TM-1. Floral apices from CCRI 36 and TM-1 were harvested and named as floral apices T and C respectively. Error bars represent standard deviation (SD).</p

Flowering time under long-day conditions, as measured by days to flowering and number of basal rosette and cauline leaves.

<p>Asterisks indicate significant variation differences between the wild-type and each <i>35S::GhFPF1</i> transgenic population line as determined by one-way ANOVA analysis. Means with SD from twenty plants of every line were shown.</p

Statistics of hypocotyl and petiole lengths of wild and transgenic plants grown under long-day conditions with fluorescent lamps.

<p>Hypocotyl lengths of wild and transgenic lines were quantified from thirty ten-day-old plants. Petiole lengths of true leaves of the basal rosette were measured and the data were gathered from thirty wild type and transgenic 24-day-old plants, respectively. Mean values (±SD) were shown and statistical analysis was evaluated by the same way as <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0091869#pone-0091869-t002" target="_blank">table 2</a>.</p

Pair-wise alignment between <i>G. raimondii</i> L. and <i>G. arboreum</i> L. orthologous sequences on amino acid and nucleic acid levels.

<p>Pair-wise alignment between <i>G. raimondii</i> L. and <i>G. arboreum</i> L. orthologous sequences on amino acid and nucleic acid levels.</p

Over-expression of <i>GhFPF1</i> led to shade-avoidance responses in transgenic plants.

<p>A. Phenotype of hypocotyl of ten-day-old wild type (left) and transgenic plants (right) grown under long-day conditions. B. The seventh rosette leaves of wild type (left) and transgenic (right) 24-day-old plants grown under the same conditions as (A). C. Shade-avoidance responses in 24-day-old transgenic plants (right) which grew fast with upward leaves. All <i>Arabidopsis</i> plants were grown under long-day conditions with high red/far red (R/FR ratio: 4.5) light provided by fluorescent lamps. D. Chlorophyll content of leaves in transgenic plants was lower than that in wild-type and the difference was very significant (P<0.01) assessed by T-test. E. The transcript levels of <i>AtPHYB</i> (AT2G18790) in wild-type and <i>GhFPF1</i> over-expression transgenic plants. The <i>AtUBQ5</i> gene was used as calibrator.</p

Multiple sequence alignment of FPF1 protein family from <i>G. hirsutum</i> L., and other species.

<p>A. Multiple alignment of FPF1 protein sequences in several species. AtFPF1 (Y11988) and AtFLP1 (AL353995) are Arabidopsis thaliana genes; SaFPF1 (Y11987), NtFPF1 (AY496934), ZmFPF1 (ACG44143) and OsRAA1 (AY659938) are from <i>Sinapis alba</i>, <i>Nicotiana tabacum</i>, <i>Zea mays</i> and <i>Oryza sativa</i>. B. Phylogenetic tree of the FPF1 proteins in the above plants as determined by the MEGA 4.1 software package.</p