Gene expression profiling of papaya (Carica papaya L.) immune response induced by CTS-N after inoculating PLDMV.

Plant immune regulation is a defensive strategy of plants for protection against pathogen invasion, and Chitosan-N (CTS-N) can induce plant autoimmunity regulation mechanisms. CTS-N was found to induce an immunomodulatory response in papaya against Papaya leaf-distortion mosaic virus (PLDMV). To date, the gene expression profile of CTS-N-induced papaya immunomodulatory response has not been reported. Here, the transcriptional map of papaya leaf genes were subjected to three treatments, viz., non-viral inoculation without CTS-N treatment (CK), virus inoculation without CTS-N treatment (CG), and virus inoculation of 1 g/L treatment (B). These were studied by pot culture experiment. Comparison of the B group with the CG group revealed 732 upregulated and 510 downregulated genes. Comparison of the CG group with the CK group revealed 909 upregulated and 1024 downregulated genes. To determine gene function, gene ontology (GO) analysis was performed, where 480 biological process genes, 256 molecular function genes, and 343 cell composition genes were differentially expressed. Kyoto Encyclopedia of Genes and Genomes (KEGG) results revealed that the top three pathways were phenylpropane biosynthesis, starch and sucrose metabolism, and plant hormone signal transduction. Real-time Quantitative PCR (qPCR) results were consistent with the transcriptome results, with a correlation coefficient of 0.87. The results of the transcriptional group showed that genes associated with plant resistance were induced by CTS-N-treatment in papaya. The chitinase gene was related to the plant disease process. Related genes in plant hormone signal transduction pathways are associated with plant resistance, and six differentially expressed genes were correlated with enhanced immune resistance in papaya.

Transgenic papaya: a useful platform for oral vaccines.

MAIN CONCLUSION: Transgenic papaya callus lines expressing the components of the S3Pvac vaccine constitute a stable platform to produce an oral vaccine against cysticercosis caused by Taenia solium or T. crassiceps. The development of effective delivery systems to cope with the reduced immunogenicity of new subunit vaccines is a priority in vaccinology. Herein, experimental evidence supporting a papaya-based platform to produce needle-free, recombinant, highly immunogenic vaccines is shown. Papaya (Carica papaya) callus lines were previously engineered by particle bombardment to express the three protective peptides of the S3Pvac anti-cysticercosis vaccine (KETc7, KETc12, KETc1). Calli were propagated in vitro, and a stable integration and expression of the target genes has been maintained, as confirmed by PCR, qRT-PCR, and HPLC. These results point papaya calli as a suitable platform for long-term transgenic expression of the vaccine peptides. The previously demonstrated protective immunogenic efficacy of S3Pvac-papaya orally administered to mice is herein confirmed in a wider dose-range and formulated with different delivery vehicles, adequate for oral vaccination. This protection is accompanied by an increase in anti-S3Pvac antibody titers and a delayed hypersensitivity response against the vaccine. A significant increase in CD4+ and CD8+ lymphocyte proliferation was induced in vitro by each vaccine peptide in mice immunized with the lowest dose of S3Pvac papaya (0.56 ng of the three peptides in 0.1 µg of papaya callus total protein per mouse). In pigs, the obliged intermediate host for Taenia solium, S3Pvac papaya was also immunogenic when orally administered in a two-log dose range. Vaccinated pigs significantly increased anti-vaccine antibodies and mononuclear cell proliferation. Overall, the oral immunogenicity of this stable S3Pvac-papaya vaccine in mice and pigs, not requiring additional adjuvants, supports the interest in papaya callus as a useful platform for plant-based vaccines.

Anti-inflammatory and immunomodulatory properties of Carica papaya.

Chronic inflammation is linked with the generation and progression of various diseases such as cancer, diabetes and atherosclerosis, and anti-inflammatory drugs therefore have the potential to assist in the treatment of these conditions. Carica papaya is a tropical plant that is traditionally used in the treatment of various ailments including inflammatory conditions. A literature search was conducted by using the keywords "papaya", "anti-inflammatory and inflammation" and "immunomodulation and immune" along with cross-referencing. Both in vitro and in vivo investigation studies were included. This is a review of all studies published since 2000 on the anti-inflammatory activity of papaya extracts and their effects on various immune-inflammatory mediators. Studies on the anti-inflammatory activities of recognized phytochemicals present in papaya are also included. Although in vitro and in vivo studies have shown that papaya extracts and papaya-associated phytochemicals possess anti-inflammatory and immunomodulatory properties, clinical studies are lacking.

Subchronic Immunotoxicity Assessment of Genetically Modified Virus-Resistant Papaya in Rats.

Papaya is an important fruit that provides a variety of vitamins with nutritional value and also holds some pharmacological properties, including immunomodulation. Genetically modified (GM) papaya plants resistant to Papaya ringspot virus (PRSV) infection have been generated by cloning the coat protein gene of the PRSV which can be used as a valuable strategy to fight PRSV infection and to increase papaya production. In order to assess the safety of GM papaya as a food, this subchronic study was conducted to assess the immunomodulatory responses of the GM papaya line 823-2210, when compared with its parent plant of non-GM papaya, Tainung-2 (TN-2), in Sprague-Dawley (SD) rats. Both non-GM and GM 823-2210 papaya fruits at low (1 g/kg bw) and high (2 g/kg bw) dosages were administered via daily oral gavage to male and female rats consecutively for 90 days. Immunophenotyping, mitogen-induced splenic cell proliferation, antigen-specific antibody response, and histopathology of the spleen and thymus were evaluated at the end of the experiment. Results of immunotoxicity assays revealed no consistent difference between rats fed for 90 days with GM 823-2210 papaya fruits, as opposed to those fed non-GM TN-2 papaya fruits, suggesting that with regard to immunomodulatory responses, GM 823-2210 papaya fruits maintain substantial equivalence to fruits of their non-GM TN-2 parent.

A current overview of the Papaya meleira virus, an unusual plant virus.

Papaya meleira virus (PMeV) is the causal agent of papaya sticky disease, which is characterized by a spontaneous exudation of fluid and aqueous latex from the papaya fruit and leaves. The latex oxidizes after atmospheric exposure, resulting in a sticky feature on the fruit from which the name of the disease originates. PMeV is an isometric virus particle with a double-stranded RNA (dsRNA) genome of approximately 12 Kb. Unusual for a plant virus, PMeV particles are localized on and linked to the polymers present in the latex. The ability of the PMeV to inhabit such a hostile environment demonstrates an intriguing interaction of the virus with the papaya. A hypersensitivity response is triggered against PMeV infection, and there is a reduction in the proteolytic activity of papaya latex during sticky disease. In papaya leaf tissues, stress responsive proteins, mostly calreticulin and proteasome-related proteins, are up regulated and proteins related to metabolism are down-regulated. Additionally, PMeV modifies the transcription of several miRNAs involved in the modulation of genes related to the ubiquitin-proteasome system. Until now, no PMeV resistant papaya genotype has been identified and roguing is the only viral control strategy available. However, a single inoculation of papaya plants with PMeV dsRNA delayed the progress of viral infection.

Genetic diversity analysis in a set of Caricaceae accessions using resistance gene analogues.

BACKGROUND: In order to assess genetic diversity of a set of 41 Caricaceae accessions, this study used 34 primer pairs designed from the conserved domains of bacterial leaf blight resistance genes from rice, in a PCR based approach, to identify and analyse resistance gene analogues from various accessions of Carica papaya, Vasconcellea goudotiana, V. microcarpa, V. parviflora, V. pubescens, V. stipulata and, V. quercifolia and Jacaratia spinosa. RESULTS: Of the 34 primer pairs fourteen gave amplification products. A total of 115 alleles were identified from 41 accesions along with 12 rare and 11 null alleles. The number of alleles per primer pair ranged from 4 to 10 with an average of 8.21 alleles/ primer pair. The average polymorphism information content value was 0.75/primer. The primers for the gene Xa1 did not give any amplification product. As a group, the Indian Carica papaya accessions produced a total of 102 alleles from 27 accessions. The similarity among the 41 accessions ranged from 1% to 53%. The dendrogram made from Jaccard's genetic similarity coefficient generated two major clusters showing that the alleles of Jacaratia spinosa and Vasconcellea accessions were distinctly different from those of Carica papaya accessions. All the alleles were sequenced and eleven of them were allotted accession numbers by NCBI. Homology searches identified similarity to rice BLB resistance genes and pseudogenes. Conserved domain searches identified gamma subunit of transcription initiation factor IIA (TFIIA), cytochrome P450, signaling domain of methyl-accepting chemotaxis protein (MCP), Nickel hydrogenase and leucine rich repeats (LRR) within the sequenced RGAs. CONCLUSIONS: The RGA profiles produced by the 14 primer pairs generated high genetic diversity. The RGA profiles identified each of the 41 accessions clearly unequivocally. Most of the DNA sequences of the amplified RGAs from this set of 41 accessions showed significant homology to the conserved regions of rice bacterial leaf blight resistance genes. These information can be used in future for large scale investigation of tentative disease resistance genes of Carica papaya and other Caricaceae genus specially Vasconcellea. Inoculation studies will be necessary to link the identified sequences to disease resistance or susceptibility.

Resistance of non-transgenic papaya plants to papaya ringspot virus (PRSV) mediated by intron-containing hairpin dsRNAs expressed in bacteria.

RNA-mediated virus resistance based on natural antiviral RNA silencing has been exploited as a powerful tool for engineering virus resistance in plants. In this study, a conserved 3'-region (positions 9839-10117, 279 nt) of the capsid protein (CP) gene of papaya ringspot virus (PRSV), designated CP279, was used to generate an intron-containing hairpin RNA (ihpRNA) construct by one-step, zero-background ligation-independent cloning (OZ-LIC). The RNaseIII-deficient Escherichia coli strain M-JM109lacY was identified as the best choice for producing large quantities of specific ihpRNA-CP279. Resistance analyses and ELISA data verified that most papaya plants mechanically co-inoculated with TRIzol-extracted ihpRNA-CP279 and PRSV were resistant to PRSV, and resistance was maintained throughout the test period (>2 months post-inoculation). In contrast, a 1-2 day interval between sequential inoculation of PRSV and ihpRNA-CP279 did not result in complete protection against PRSV infection, but delayed the appearance of viral symptoms by 3 to 4 days. These findings indicate that direct mechanical inoculation of papaya plants with bacterially-expressed ihpRNA-CP279 targeting the PRSV CP gene can interfere with virus infection. This work lays a foundation for developing a non-transgenic approach to control PRSV by directly spraying plants with ihpRNA or crude bacterial extract preparations.

Efficiency of circulant diallels via mixed models in the selection of papaya genotypes resistant to foliar fungal diseases.

Diallel crossing methods provide information regarding the performance of genitors between themselves and their hybrid combinations. However, with a large number of parents, the number of hybrid combinations that can be obtained and evaluated become limited. One option regarding the number of parents involved is the adoption of circulant diallels. However, information is lacking regarding diallel analysis using mixed models. This study aimed to evaluate the efficacy of the method of linear mixed models to estimate, for variable resistance to foliar fungal diseases, components of general and specific combining ability in a circulant table with different s values. Subsequently, 50 diallels were simulated for each s value, and the correlations and estimates of the combining abilities of the different diallel combinations were analyzed. The circulant diallel method using mixed modeling was effective in the classification of genitors regarding their combining abilities relative to the complete diallels. The numbers of crosses in which each genitor(s) will compose the circulant diallel and the estimated heritability affect the combining ability estimates. With three crosses per parent, it is possible to obtain good concordance (correlation above 0.8) between the combining ability estimates.

[Anaphylaxis secondary to prick-to-prick tests to foods and its risk factors]. / Anafilaxia secundaria a pruebas cutáneas prick-to-prick para alimentos y sus factores de riesgo.

The diagnosis of food allergy requires a proper anamnesis and diagnostic testing with skin prick tests with fresh foods and/or standardized allergen, or specific IgE tests. The risk of systemic reactions is of 15-23 per 100,000 skin tests performed by prick method, specifically anaphylaxis at 0.02%. This paper reports the case of four patients, who while performing prick to prick test with fresh food presented anaphylactic reaction. Implicated foods were fruits of the Rosaceae, Anacardiaceae and Caricaceae families. The severity of anaphylaxis was: two patients with grade 4, one patient grade 2 and one grade 3, all with appropriate response to drug treatment. The risk factors identified were: female sex, personal history of atopy, previous systemic reaction to Hymenoptera venom, prior anaphylaxis to prick tests to aeroallergens. We found that a history of positive skin test for Betulla v, can be a risk factor for anaphylaxis in patients with oral syndrome. During testing prick to prick with food anaphylaxis can occur, so it should be made with aerial red team on hand. The history of positivity Betulla v is an additional risk factor in these patients.

El diagnóstico de alergia alimentaria requiere una anamnesis adecuada y la realización de pruebas diagnósticas, las pruebas cutáneas con alimentos en fresco, con alergenos estandarizados, o con ambos; las pruebas de IgE específica para alimentos son útiles. El riesgo de reacciones sistémicas por pruebas cutáneas por punción es de 15 a 23 por cada 100,000 y el de anafilaxia es de 0.02%. Comunicamos el caso de cuatro pacientes que sufrieron anafilaxia durante la realización de prueba Prick-to-Prick con alimentos frescos. Los alimentos implicados fueron frutas de las familias Rosaceae, Anacardiaceae y Caricaceae. En dos pacientes la anafilaxia fue de grado 4, en una grado 2 y en otra grado 3, todas con adecuada respuesta al tratamiento farmacológico. Los factores de riesgo fueron: sexo femenino, antecedente personal de atopia, reacción sistémica previa a veneno de himenópteros y anafilaxia previa con pruebas por punción para aeroalergenos. Las cuatro pacientes tuvieron síndrome de alergia oral y 50% tenía antecedente de prueba por punción positiva a betuláceas. Durante la realización de pruebas Prick-to-Prick para alimentos los pacientes pueden presentar anafilaxia, por lo que deben realizarse en un área que cuente con equipo rojo. El antecedente de síndrome de alergia oral se observó en todos los casos y la mitad de las pacientes tuvieron positividad a betuláceas, estos antecedentes pueden ser factores de riesgo adicional de anafilaxia dura te la realización de pruebas Prick-to-Prick para alimentos.

Complete genome sequence of an isolate of papaya leaf distortion mosaic virus from commercialized PRSV-resistant transgenic papaya in China.

Papaya leaf distortion mosaic virus is highly destructive to commercial papaya production. Here, the complete genome sequence was determined for an isolate of papaya leaf distortion mosaic virus, designated PLDMV-DF, infecting the commercialized papaya ringspot virus (PRSV)-resistant transgenic papaya from China. Excluding the 3'-poly (A) tail, the sequence shares high sequence identity to several PLDMV isolates from Taiwan and Japan and is phylogenetically most closely related to the isolate from Japan. Infection of PLDMV-DF in transgenic PRSV-resistant papaya may indicate emergence of this disease in genetically engineered plants. The reported sequence for this isolate may help generate bi-transgenic papaya resistant to PRSV and PLDMV.

A set of host proteins interacting with papaya ringspot virus NIa-Pro protein identified in a yeast two-hybrid system.

UNLABELLED: The protein-protein interactions between viral and host proteins play an essential role in plant virus infection and host defense. The potyviral nuclear inclusion protein a protease (NIa-Pro) is involved in various steps of viral infection. In this study, the host proteins interacting with papaya ringspot virus (PRSV) NIa-Pro were screened in a Carica papaya L. plant cDNA library using a Sos recruitment two-hybrid system (SRS). We confirmed that the full-length EIF3G, FBPA1, FK506BP, GTPBP, MSRB1, and MTL from papaya can interact specifically with PRSV NIa-Pro in yeast, respectively. These proteins fufill important functions in plant protein translation, biotic and abiotic stress, energy metabolism and signal transduction. In this paper, we discuss possible functions of interactions between these host proteins and NIa-Pro in PRSV infection and their role in host defense. KEYWORDS: Sos recruitment two-hybrid system; papaya ringspot virus; NIa-Pro; protein-protein interaction.

Identification and detection method for genetically modified papaya resistant to papaya ringspot virus YK strain.

Unauthorized genetically modified (GM) papaya (Carica papaya LINNAEUS) was detected in a commercially processed product, which included papaya as a major ingredient, in Japan. We identified the transgenic vector construct generated based on resistance to infection with the papaya ringspot virus (PRSV) YK strain. A specific detection method to qualitatively monitor papaya products for contamination with the GM papaya was developed using the real-time polymerase chain reaction.

Allergenicity assessment of the papaya ringspot virus coat protein expressed in transgenic rainbow papaya.

The virus-resistant, transgenic commercial papaya Rainbow and SunUp (Carica papaya L.) have been consumed locally in Hawaii and elsewhere in the mainland United States and Canada since their release to planters in Hawaii in 1998. These papaya are derived from transgenic papaya line 55-1 and carry the coat protein (CP) gene of papaya ringspot virus (PRSV). The PRSV CP was evaluated for potential allergenicity, an important component in assessing the safety of food derived from transgenic plants. The transgene PRSV CP sequence of Rainbow papaya did not exhibit greater than 35% amino acid sequence homology to known allergens, nor did it have a stretch of eight amino acids found in known allergens which are known common bioinformatic methods used for assessing similarity to allergen proteins. PRSV CP was also tested for stability in simulated gastric fluid and simulated intestinal fluid and under various heat treatments. The results showed that PRSV CP was degraded under conditions for which allergenic proteins relative to nonallergens are purported to be stable. The potential human intake of transgene-derived PRSV CP was assessed by measuring CP levels in Rainbow and SunUp along with estimating the fruit consumption rates and was compared to potential intake estimates of PRSV CP from naturally infected nontransgenic papaya. Following accepted allergenicity assessment criteria, our results show that the transgene-derived PRSV CP does not pose a risk of food allergy.

Strategy for a generic resistance to geminiviruses infecting tomato and papaya through in silico siRNA search.

Use of siRNA is a powerful methodology to particularly knockdown the targeted genes in a sequence specific manner. The potential of siRNA can be harnessed for silencing specific geminiviral genes in papaya and tomato plant hosts, thus making them resistant to the respective viruses. The challenge is in designing exogenous siRNA which can trigger silencing of viral genes irrespective of the genetic variability in different viral isolates and at the same time the selected siRNA does not target any plant gene (off target silencing). In this study, we have designed siRNA from the most conserved regions of viral coat protein (AV1) and replicase (AC1) genes retrieved from different isolates of geminiviruses infecting papaya (PLCV), and tomato (TLCV & TLCV, Northern India), so as to give a broad spectrum resistance and efficient silencing as it is highly homology-dependent strategy. Software siRNA finder (Ambion) was used on the selected conserved sequences in order to select only those putative siRNA oligonucleotides which fulfill all the basic criteria required as per the algorithm. Finally, a cross search using BLAST was performed to confirm that the designed siRNAs do not have any homology to plant genome sequences. The putative siRNA sequences thus designed to target essential genes of geminiviruses and introduced into the plants may facilitate developing papaya and tomato crops with generic resistance to geminiviruses.

Cucumber anaphylaxis in a latex-sensitized patient.

We report the case of a 76-year-old woman who experienced dizziness, vomiting, dyspnea, thoracic erythema, and vaginal itching within 5 minutes of eating cucumber. She had been diagnosed 3 months earlier with papaya urticaria and latex sensitization. The results of skin prick tests were positive for cucumber, watermelon, papaya, and latex and negative for melon and profilin extracts. ImmunoCAP for latex-specific serum immunoglobulin (Ig) E was positive. Cucumber-specific serum IgE was negative. Immunoblot analysis using patient serum revealed a 30- to 32-kDa protein band in the cucumber (peel) and papaya extracts. Immunoblot inhibition with latex extract demonstrated inhibition of the band in both extracts. Immunoblot inhibition with cucumber-papaya and papaya-cucumber revealed inhibition of the same band in the cucumber and papaya extracts, respectively. We present a case of IgE-mediated allergy to cucumber and papaya. Our results strongly suggest that the allergen(s) implicated are associated with latex sensitization. To our knowledge, this is the first report of cucumber-latex and cucumber-papaya cross-reactivity.

Generation of hermaphrodite transgenic papaya lines with virus resistance via transformation of somatic embryos derived from adventitious roots of in vitro shoots.

Papaya production is seriously limited by Papaya ringspot virus (PRSV) worldwide and Papaya leaf-distortion mosaic virus (PLDMV) in Eastern Asia. An efficient transformation method for developing papaya lines with transgenic resistance to these viruses and commercially desirable traits, such as hermaphroditism, is crucial to shorten the breeding program for this fruit crop. In this investigation, an untranslatable chimeric construct pYP08 containing truncated PRSV coat protein (CP) and PLDMV CP genes coupled with the 3' untranslational region of PLDMV, was generated. Root segments from different portions of adventitious roots of in vitro multiple shoots of hermaphroditic plants of papaya cultivars 'Tainung No. 2', 'Sunrise', and 'Thailand' were cultured on induction medium for regeneration into somatic embryos. The highest frequency of somatic embryogenesis was from the root-tip segments of adventitious roots developed 2-4 weeks after rooting in perlite medium. After proliferation, embryogenic tissues derived from somatic embryos were wounded in liquid-phase by carborundum and transformed by Agrobacterium carrying pYP08. Similarly, another construct pBG-PLDMVstop containing untranslatable CP gene of PLDMV was also transferred to 'Sunrise' and 'Thailand', the parental cultivars of 'Tainung No. 2'. Among 107 transgenic lines regenerated from 349 root-tip segments, nine lines of Tainung No. 2 carrying YP08 were highly resistant to PRSV and PLDMV, and 9 lines (8 'Sunrise' and 1 'Thailand') carrying PLDMV CP highly resistant to PLDMV, by a mechanism of post-transcriptional gene silencing. The hermaphroditic characteristics of the transgenic lines were confirmed by PCR with sex-linked primers and phenotypes of flower and fruit. Our approach has generated transgenic resistance to both PRSV and PLDMV with commercially desirable characters and can significantly shorten the time-consuming breeding programs for the generation of elite cultivars of papaya hybrids.

Generation of transgenic papaya with double resistance to Papaya ringspot virus and Papaya leaf-distortion mosaic virus.

During the field tests of coat protein (CP)-transgenic papaya lines resistant to Papaya ringspot virus (PRSV), another Potyvirus sp., Papaya leaf-distortion mosaic virus (PLDMV), appeared as an emerging threat to the transgenic papaya. In this investigation, an untranslatable chimeric construct containing the truncated CP coding region of the PLDMV P-TW-WF isolate and the truncated CP coding region with the complete 3' untranslated region of PRSV YK isolate was transferred into papaya (Carica papaya cv. Thailand) via Agrobacterium-mediated transformation to generate transgenic plants with resistance to PLDMV and PRSV. Seventy-five transgenic lines were obtained and challenged with PRSV YK or PLDMV P-TW-WF by mechanical inoculation under greenhouse conditions. Thirty-eight transgenic lines showing no symptoms 1 month after inoculation were regarded as highly resistant lines. Southern and Northern analyses revealed that four weakly resistant lines have one or two inserts of the construct and accumulate detectable amounts of transgene transcript, whereas nine resistant lines contain two or three inserts without significant accumulation of transgene transcript. The results indicated that double virus resistance in transgenic lines resulted from double or more copies of the insert through the mechanism of RNA-mediated posttranscriptional gene silencing. Furthermore, three of nine resistant lines showed high levels of resistance to heterologous PRSV strains originating from Hawaii, Thailand, and Mexico. Our transgenic lines have great potential for controlling a number of PRSV strains and PLDMV in Taiwan and elsewhere.

Transgene-specific and event-specific molecular markers for characterization of transgenic papaya lines resistant to Papaya ringspot virus.

The commercially valuable transgenic papaya lines carrying the coat protein (CP) gene of Papaya ringspot virus (PRSV) and conferring virus resistance have been developed in Hawaii and Taiwan in the past decade. Prompt and sensitive protocols for transgene-specific and event-specific detections are essential for traceability of these lines to fulfill regulatory requirement in EU and some Asian countries. Here, based on polymerase chain reaction (PCR) approaches, we demonstrated different detection protocols for characterization of PRSV CP-transgenic papaya lines. Transgene-specific products were amplified using different specific primer pairs targeting the sequences of the promoter, the terminator, the selection marker, and the transgene, and the region across the promoter and transgene. Moreover, after cloning and sequencing the DNA fragments amplified by adaptor ligation-PCR, the junctions between plant genomic DNA and the T-DNA insert were elucidated. The event-specific method targeting the flanking sequences and the transgene was developed for identification of a specific transgenic line. The PCR patterns using primers designed from the left or the right flanking DNA sequence of the transgene insert in three selected transgenic papaya lines were specific and reproducible. Our results also verified that PRSV CP transgene is integrated into transgenic papaya genome in different loci. The copy number of inserted T-DNA was further confirmed by real-time PCR. The event-specific molecular markers developed in this investigation are crucial for regulatory requirement in some countries and intellectual protection. Also, these markers are helpful for prompt screening of a homozygote-transgenic progeny in the breeding program.

Research advances on transgenic plant vaccines.

In recent years, with the development of genetics molecular biology and plant biotechnology, the vaccination (e.g. genetic engineering subunit vaccine, living vector vaccine, nucleic acid vaccine) programs are taking on a prosperous evolvement. In particular, the technology of the use of transgenic plants to produce human or animal therapeutic vaccines receives increasing attention. Expressing vaccine candidates in vegetables and fruits open up a new avenue for producing oral/edible vaccines. Transgenic plant vaccine disquisitions exhibit a tempting latent exploiting foreground. There are a lot of advantages for transgenic plant vaccines, such as low cost, easiness of storage, and convenient immune-inoculation. Some productions converged in edible tissues, so they can be consumed directly without isolation and purification. Up to now, many transgenic plant vaccine productions have been investigated and developed. In this review, recent advances on plant-derived recombinant protein expression systems, infectious targets, and delivery systems are presented. Some issues of high concern such as biosafety and public health are also discussed. Special attention is given to the prospects and limitations on transgenic plant vaccines.