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SUMMARY: The Izu Islands of southcentral Japan are thought to fall within the distribution range of Neoporphyra dentata. However, the gametophytic blades of Bangiales collected from Shikinejima and Hachijojima, Izu Islands, were identified as Neoporphyra haitanensis in our previous study. Thus, it became uncertain whether N. dentata is distributed in the Izu Islands, including Shikinejima. To clarify whether N. dentata grows on Shikinejima, we conducted a further distribution survey of N. dentata on the island. The morphological features of the blade samples collected from an additional sampling site on Shikinejima were more similar to those of N. dentata than to those of N. haitanensis: the blade thickness and the division formula of spermatangia resembled those of the former species rather than the latter species. However, the division formula of zygotosporangia was different from those of either species. The phylogenetic analyses of the rbcL gene indicated that the samples were resolved in a clade including N. dentata collected from Shirahama, Chiba Prefecture, and Enoshima, Kanagawa Prefecture, Honshu, Japan. The p‐distances of the chloroplast rbcL gene and nuclear 18S rRNA also supported identification of the samples as N. dentata. The results demonstrated that N. dentata is also distributed on Shikinejima with co‐occurring N. haitanensis, and that the island materials of the two species are genetically different from other materials of the two species, respectively. [ABSTRACT FROM AUTHOR]

We conducted preliminary experimental cultivation using the OHN-1 strain of Neoporphyra kitoi to confirm the growth of the gametophytic blades in the nori farm. The mature and immature long linear blades grew in the nori farm and were confirmed to be N. kitoi based on the nucleotide sequences of the chloroplast RUBISCO spacer region. Furthermore, we developed PCR–RFLP analyses using chloroplast and nuclear DNA to discriminate simply between N. kitoi and Neopyropia yezoensis, the species cultivated in Japan, for identifying many samples for growth comparisons between the two species during future experimental cultivation. In addition, four major FAA contents [alanine (Ala), aspartic acid (Asp), glutamic acid (Glu) and taurine (Tau)] were compared between the blades of N. kitoi OHN-1 and cultivated Neopy. yezoensis (Neopyropia yezoensis f. narawaensis) HG-4 under identical laboratory conditions. As a result, the contents of the four major FAAs, especially Ala, were significantly higher in OHN-1 than in HG-4. The results suggested that the OHN-1 strain of N. kitoi has potential as a new cultivated species with high growth and a better flavor. [ABSTRACT FROM AUTHOR]

To confirm the distribution of the allopolyploid and ancestral species in the Matsukawaura Lagoon in a hybrid zone between Neopyropia yezoensis and Neopyropia tenera, molecular marker analyses and culture experiments were performed using collected blades. All the blades examined, except for one blade identified as N. tenera, exhibited nuclear genotypes of both species, but only the chloroplast haplotype of N. yezoensis. In addition, four conchocelis strains were isolated from each of the blades, and they contained genotypes of both N. yezoensis and N. tenera; a conchocelis strain from the sole N. tenera blade was also isolated. Using the conchocelis strains, F1 blades that were produced from archeospores were obtained under laboratory culture and examined by nuclear DNA markers. These results indicated that the collected blades and the examined F1 blades, except for the single N. tenera blade, were allodiploids and that the four conchocelis strains could be allotetraploids derived from interspecific hybridization between N. yezoensis as the maternal strain and N. tenera as the paternal strain. Therefore, most of the Neopyropia blades growing in the Matsukawaura Lagoon are presumed to be from allopolyploid lineages derived from interspecific hybridization. Furthermore, a growth comparison between the allopolyploid strain and the N. tenera strain was carried out under identical culture conditions. The rate of increase in blade length was significantly higher in the allopolyploid hybrid than in the pure N. tenera. The results suggest that further exploration of allopolyploidy in the potential hybrid zones will be needed not only for the investigation of speciation mechanisms in the bladed Bangiales but also for the genetic improvement of the marine crop. [ABSTRACT FROM AUTHOR]

SUMMARY We conducted a morphological observation and molecular phylogenetic analyses on bladed Bangiales collected from natural populations on Shikinejima Island and Hachijojima Island of the Izu Islands, southern central Japan. The morphological features of the gametophytic blades such as the blade shape, blade margin and reproductive cell divisions were extremely similar to those parameters of Neoporphyra haitanensis, a species cultivated in southern China, and Neoporphyra dentata. However, the blades collected from the two islands were thicker than the blades of N. dentata; this feature was the same as that of N. haitanensis. The chloroplast RUBISCO spacer sequences of present samples were identical to the sequences of N. haitanensis from China. Moreover, phylogenetic positions of three conchocelis strains isolated from the blades on the two islands were examined based on concatenated sequences of the chloroplast rbcL gene and nuclear 18S rRNA. The results showed that the strains were resolved in a clade including N. haitanensis; however, the strains were separated from N. haitanensis in China. Therefore, the bladed Bangiales from the two islands were identified as N. haitanensis. The present study is the first record of N. haitanensis in natural populations in Japan. [ABSTRACT FROM AUTHOR]

A green-colored gametophytic blade of wild Pyropia was found in a natural population in the estuary area of the Naruse River, northeast Japan. To identify the isolated green mutant strain K-G1 and the wild-type strain K-1 derived from the same population, we carried out morphological observations and molecular analysis of the RUBISCO spacer region and the rbcL gene. As a result, the two strains were identified as Pyropia kinositae, which is a species closely related to the marine crop Pyropia yezoensis. Under the same laboratory culture, the growth and pigment contents of the green mutant K-G1 were compared with those of the wild-type K-1. The growth of the conchocelis and gametophytic blade tended to be slower in the green mutant than in the wild type, although differences in their growth were not apparent in some of the temperature conditions examined. Maturation of the conchocelis and blade in the two strains was fastest at 20 °C among the temperature conditions examined (5–25 °C). The phycoerythrin content was significantly lower in the green mutant than in the wild type. Additionally, the color phenotype of the green mutant was genetically stable in both the sporophytic conchocelis and gametophytic phases. Because the color of a pigmentation mutant facilitates cross-confirmation as a genetic marker in monoecious Pyropia species, these results suggest that the green mutant of P. kinositae will contribute as new breeding material for cross-breeding and polyploidy breeding in Pyropia. [ABSTRACT FROM AUTHOR]

While subject to illumination, photosystem I (PSI) has the potential to produce reactive oxygen species (ROS) that can cause photo-oxidative damage in oxygenic photoautotrophs. The reaction center chlorophyll in PSI (P700) is kept oxidized in excess light conditions to limit over-excitation of PSI and alleviate the production of ROS. Oxidation of P700 requires a sufficient electron sink for PSI, which is responsible for flavodiiron proteins (FLV) safely dissipating electrons to O2 in cyanobacteria, green algae, and land plants except for angiosperms during short-pulse light (SP) illumination under which photosynthesis and photorespiration do not occur. This fact implies that O2 usage is essential for P700 oxidation but also raises the question why angiosperms lost FLV. Here, we first found that aquatic photoautotrophs in red plastid lineage, in which no gene for FLV has been found, could keep P700 oxidized during SP illumination alleviating the photo-oxidative damage in PSI even without O2 usage. We comprehensively assessed P700 oxidation during SP illumination in the presence and absence of O2 in cyanobacteria (Cyanophyta), green algae (Chlorophyta), angiosperms (Streptophyta), red algae (Rhodophyta), and secondary algae (Cryptophyta, Haptophyta, and Heterokontophyta). A variety of dependencies of P700 oxidation on O2 among these photoautotrophs clearly suggest that O2 usage and FLV are not universally required to oxidize P700 for protecting PSI against ROS damage. Our results expand the understanding of the diverse strategies taken by oxygenic photoautotrophs to oxidize P700 and mitigate the risks of ROS. [ABSTRACT FROM AUTHOR]

In the marine crop Pyropia yezoensis, a cross-fertilization experiment between a cultivated strain and cryptic species was carried out to investigate the possibility of polyploidy breeding using cryptic species. To perform the cross-fertilization, we isolated a green mutant strain (IBYC-G1) of the cryptic species by heavy-ion beam irradiation. The green mutant was crossed with a wildtype strain (HG-4) of P. yezoensis f. narawaensis. The F1 heterozygous conchocelis developed normally and released numerous conchospores. However, almost all the conchospore germlings did not survive past the four-cell stage or thereabout, and a few germlings developed into gametophytic blades. These results indicated that reproductive isolation occurred during meiosis and that P. yezoensis and cryptic species were different species using the biological species concept. Almost all the archeospores that were released from the surviving F1 blades developed into normal gametophytic blades. The result of nuclear DNA marker analysis revealed that the F1 blades that developed from both conchospores and archeospores were allodiploids in the haploid gametophytic phase. Although their F1 blades were extremely slender, their blade color was a deeper reddish brown as compared with that of P. yezoensis f. narawaensis, and the rate of blade length increase was remarkably higher in the blades of F1 than in those of the cryptic species. These results suggested that polyploidy breeding in the marine crop will attract more attention as a breeding method for further development of new cultivars. [ABSTRACT FROM AUTHOR]

The light-harvesting antennas of oxygenic photosynthetic organisms capture light energy and transfer it to the reaction centers of their photosystems. The light-harvesting antennas of cyanobacteria and red algae, called phycobilisomes (PBSs), supply light energy to both photosystem I (PSI) and photosystem II (PSII). However, the excitation energy transfer processes from PBS to PSI and PSII are not understood in detail. In the present study, the energy transfer processes from PBS to PSs in various cyanobacteria and red algae were examined in vivo by selectively exciting their PSs or PBSs, and measuring the resulting picosecond to nanosecond time-resolved fluorescences. By observing the delayed fluorescence spectrum of PBS-selective excitation in Arthrospira platensis, we demonstrated that energy transfer from PBS to PSI via PSII (PBS→PSII→PSI transfer) occurs even for PSI trimers. The contribution of PBS→PSII→PSI transfer was species dependent, being largest in the wild-type of red alga Pyropia yezoensis (formerly Porphyra yezoensis) and smallest in Synechococcus sp. PCC 7002. Comparing the time-resolved fluorescence after PSs- and PBS-selective excitation, we revealed that light energy flows from CP43 to CP47 by energy transfer between the neighboring PSII monomers in PBS-PSII supercomplexes. We also suggest two pathways of energy transfer: direct energy transfer from PBS to PSI (PBS→PSI transfer) and indirect transfer through PSII (PBS→PSII→PSI transfer). We also infer that PBS→PSI transfer conveys light energy to a lower-energy red chlorophyll than PBS→PSII→PSI transfer. [ABSTRACT FROM AUTHOR]

To clarify the significance of genetic resources in the marine crop Undaria pinnatifida, we investigated whether morphological features of two cultivars were genetically different from each other based on combined studies of morphological and molecular analyses. The two cultivars were produced, respectively, by self-fertilization between male and female gametophytes, which were each developed from a single zoospore from two morphologically different sporophytes cultivated in the eastern Seto Inland Sea, Japan. Their sporophytes, each with the same genotype, were cultivated in an Undaria farm under the same environmental conditions and were periodically collected for morphometric comparisons. The results of this experimental cultivation demonstrate that various morphological features, such as the blade, stipe, and sporophyll shapes, and developmental processes of the divided blade and sporophyll, clearly differed between the two cultivars. Although the two cultivars showed identical haplotypes in both of the mitochondrial cox3 gene and the tatC- tLeu region, they exhibited different genotypes at the microsatellite loci. These results indicate that morphological and developmental differences were genetically controlled, in addition to phenotypic plasticity due to changes in environmental conditions. It is therefore necessary for the marine crop breeding program to collect more mature sporophytes of cultivated and wild U. pinnatifida from various regions and to establish more female and male gametophyte strains each developed from a single zoospore. The origin of U. pinnatifida cultivated in the eastern Seto Inland Sea, famous for Japanese Undaria cultivation, is also discussed. [ABSTRACT FROM AUTHOR]

Understanding the nutrient uptake kinetics of kelp populations will contribute to an improved understanding of environmental adaptation and the breeding of new cultivars. In this study, we examined the morphological characteristics, carbon (C) and nitrogen (N) contents, and NO -N and NH -N uptake kinetics of Undaria pinnatifida sporophytes cultivated at six industrial farms throughout Japan. We detected significant differences in morphology among sites. At Matsushima Bay (northern Pacific coast of Japan), where autumnal seawater temperatures fall in concert with increasing nutrient concentrations, sporophytes were significantly larger than at other sites from December to February. The C content of the sporophytes was seasonally stable at all of the locations, but the N content of sporophytes declined after February due to a decrease in seawater nutrients. We compared the uptake kinetics of NO -N and NH -N among cultivation sites. V and K , which are Michaelis-Menten parameters that measure adaptation to nutrient concentrations, were highest in the Seto Inland Sea and lowest in the northern sector of the Sea of Japan. The V / K ratio is a measure of adaptation to low nutrient concentrations; the highest values were measured in the northern sector of the Sea of Japan. The parameter ranges were broader than those previously reported for invasive populations of U. pinnatifida in other parts of the world. Thus, we detected population-level adaptations to the various nutrient conditions in Japanese waters, and these results suggest the existence of ecotypes according to nutrient uptake kinetics. The different populations may be used to provide sources of genetic material that could be of value in breeding programmes by improving productivity and quality. [ABSTRACT FROM AUTHOR]

In the marine red alga Pyropia yezoensis, commonly known in Japan as nori, sympatric occurrence of two cryptic species Pyropia sp. 2 and Pyropia sp. 3 on the same rock in a natural habitat has been confirmed by molecular analysis and detailed morphological observations. To confirm whether Pyropia sp. 2 and Pyropia sp. 3 were reproductively isolated in the sympatric population, 170 blades that had previously been studied using a maternally inherited plastid marker were examined with a nuclear gene marker. The results suggested that Pyropia sp. 2 and Pyropia sp. 3 with identical morphological features were reproductively isolated in the sympatric population and that they were different species based on the biological species concept. Although gametophytic blades of Pyropia were usually assumed to be haploid, 18 of 170 blades possessed both of the two genotypes derived from Pyropia sp. 2 and from Pyropia sp. 3. These results inferred that allodiploid blades were generated from the interspecific hybridization between these two cryptic species. The present findings provide insights for future studies on the speciation mechanism in seaweeds, particularly for genera that contain numerous species. [ABSTRACT FROM AUTHOR]

In a previous study on wild populations of Pyropia, the occurrence of two possible new species ( Pyropia sp. 2 and Pyropia sp. 3) which are closely related to the two commercially important Pyropia species, P. yezoensis and P. tenera, was confirmed as the result of molecular phylogenetic analyses. To characterize the morphological features of the two wild Pyropia species, we collected Pyropia blades in a natural population in which Pyropia sp. 3 was known to occur, and carried out molecular identification before detailed morphological observations. Through the molecular identification we found, unexpectedly, that Pyropia sp. 2 blades grew sympatrically in the same site. Therefore, after molecular identification, we examined in detail the external morphology and anatomy of the two wild Pyropia species using more than 10 blades each. As a result, it is concluded that all of the blades of the two species are morphologically identical to P. yezoensis, but distinct from P. tenera. It is therefore considered that both of the two wild Pyropia species are cryptic species within the P. yezoensis complex. Furthermore, this study revealed that the two cryptic species grew sympatrically, even on the same rocks within the natural habitat. [ABSTRACT FROM AUTHOR]

In the marine crop Pyropia yezoensis (Ueda) M. S. Hwang et H. G. Choi, it is known that conchospores from heterozygous conchocelis develop into sectored gametophytic blades (chimeras), but archeospores asexually released from haploid blades do not usually grow into chimeric blades. In this study, chimeras with mosaic pattern consisting of the green and wildtype colors were developed from archeospores that were released from a blade piece containing a cell cluster of green color induced by heavy-ion beam irradiation. To make clear whether these archeospores were produced from the green-colored cells or the wildtype-colored cells, cell clusters of the green mutant, wildtype, and mosaic pattern were cut out from the grown chimera, and archeospores were released from each of the three blade pieces. Archeospores from the green-mutant blade piece and from the wildtype blade piece developed into only green-mutant blades and wildtype blades, respectively. In contrast, archeospores from the blade piece with mosaic pattern developed into green-mutant blades, wildtype blades, and chimeric blades with mosaic pattern of the two colors, although the frequency of the chimeras was low. Because each gametophytic cell possesses a single plastid, it is difficult to explain the occurrence of the new chimeras as a mutation of the plastid DNA. Thus, the new chimeras are considered to be due to transposable elements in Pyropia. [ABSTRACT FROM AUTHOR]

To confirm whether allopolyploidy occurs in samples of previously identified Porphyra yezoensis Ueda, P. tenera Kjellm., and P. yezoensis × P. tenera from natural and cultivated populations, we examined these samples by using PCR-RFLP and microsatellite analyses of multiple nuclear and chloroplast regions [nuclear regions: type II DNA topoisomerase gene ( TOP2), actin-related protein 4 gene ( ARP4), internal transcribed spacer (ITS) rDNA and three microsatellite loci; chloroplast region: RUBISCO spacer]. Except for the ITS region, these multiple nuclear markers indicated that the wild strain MT-1 and the cultivated strain 90-02 (previously identified as P. yezoensis × P. tenera and cultivated P. tenera, respectively) are heterozygous and possess both genotypes of P. tenera and P. yezoensis in the conchocelis phase. Furthermore, gametophytic blades of two pure lines, HG-TY1 and HG-TY2 (F strains of MT-1 and 90-02, respectively), were also heterozygous, and six chromosomes per single cell could be observed in each blade of the two pure lines. These results demonstrate that allopolyploidy occurs in Porphyra strains derived from both natural and cultivated populations, even though ITS genotypes of these strains showed homogenization toward one parental ITS. [ABSTRACT FROM AUTHOR]

We performed interspecific hybridization in the haploid blade-forming marine species (nori) of the genus Porphyra, which have a heteromorphic life cycle with a haploid gametophytic blade and a diploid microscopic sporophyte called the “conchocelis phase.” The green mutant HGT-6 of P. tenera var. tamatsuensis A. Miura was crossed with the wildtype HG-1 of P. yezoensis f. narawaensis A. Miura; the F1 heterozygous conchocelis developed normally and released numerous conchospores. However, almost all the conchospore germlings did not survive past the four-cell stage or thereabouts, and only a few germlings developed into gametophytic blades. These results indicate that hybrid breakdown occurred during the meiosis, while the surviving F1 gametophytic blades were considered a breakthrough in the interspecific hybridization of Porphyra. Organelle genomes (cpDNA and mtDNA) were found to be maternally inherited in the interspecific hybridization by molecular analyses of the organelle DNA. In particular, molecular analyses of nuclear DNA revealed that the surviving F1 blades were allodiploids in the haploid gametophytic phase; however, there is a possibility of the occurrence of rapid chromosomal locus elimination and rearrangement in the F1 conchocelis phase. Our findings are noteworthy to the breeding of cultivated Porphyra and will provide important information for understanding of the speciation of marine plants with high species diversity. [ABSTRACT FROM AUTHOR]

To discriminate between cultivated Porphyra species ( Porphyra yezoensis and Porphyra tenera) and closely related wild Porphyra species, we developed a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis of the rbcL gene using five restriction enzymes. Although our previous PCR-RFLP analyses of internal transcribed spacer (ITS) rDNA and plastid RuBisCO spacer regions could not always discriminate wild P. yezoensis, wild P. tenera, and closely related wild species, the PCR-RFLP profiles of the rbcL gene were useful in discriminating samples collected from natural habitats. Therefore, PCR-RFLP analysis of the rbcL gene will help in the simple identification of a large number of samples, not only for the establishment of reliable cultures as breeding material, but also for the taxonomic investigations of species that are closely related to cultivated Porphyra. [ABSTRACT FROM AUTHOR]

The present study describes the isolation of pigmentation mutants of Porphyra yezoensis Ueda induced by heavy-ion beam irradiation for the first time. The gametophytic blades were irradiated with 12C+6 ion beams within a dose range of 25–400 Gy. From the survival rate and cell growth of the irradiated blades, it is suggested that a dose of 150 Gy or less is suitable to induce mutation for the isolation of mutants of P. yezoensis. After irradiation, red, green and deep reddish brown-colored gametophytic blades developed from archeospores that were released from each of the mutated cell clusters of the respective different colors, and the red mutant strain (IBY-R1) and green mutant strain (IBY-G1) were established as a conchocelis colony in culture. Blades of the mutants were characterized by their growth and photosynthetic pigment contents compared with those of the wild-type. From these results, it is clear that heavy-ion beam mutagenesis will be an effective tool for genetic and breeding studies of Porphyra, and also for other algal research. [ABSTRACT FROM AUTHOR]

We investigated the genetic variations of the samples that were tentatively identified as two cultivated Porphyra species ( Porphyra yezoensis Ueda and Porphyra tenera Kjellm.) from various natural populations in Japan using molecular analyses of plastid and nuclear DNA. From PCR-RFLP analyses using nuclear internal transcribed spacer (ITS) rDNA and plastid RUBISCO spacer regions and phylogenetic analyses using plastid rbcL and nuclear ITS-1 rDNA sequences, our samples from natural populations of P. yezoensis and P. tenera showed remarkably higher genetic variations than found in strains that are currently used for cultivation. In addition, it is inferred that our samples contain four wild Porphyra species, and that three of the four species, containing Porphyra kinositae, are closely related to cultivated Porphyra species. Furthermore, our PCR-RFLP and molecular phylogenetic analyses using both the nuclear and plastid DNA demonstrated the occurrence of plastid introgression from P. yezoensis to P. tenera and suggested the possibility of plastid introgression from cultivated P. yezoensis to wild P. yezoensis. These results imply the importance of collecting and establishing more strains of cultivated Porphyra species and related wild species from natural populations as genetic resources for further improvement of cultivated Porphyra strains. [ABSTRACT FROM AUTHOR]

Porphyra yezoensis and P. tenera are the representative species of the marine crop Porphyra (nori) in Japan. Since the two species are extremely similar to each other in morphology, nori breeders have tentatively classified many strains of cultivated Porphyra into the two species without strict species identification. In order to facilitate rapid and reliable identification of the many strains of Porphyra currently cultivated in various Japanese regions, 24 conchocelis strains were examined by polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) analysis using the plastid RuBisCo spacer and the nuclear internal transcribed spacer regions. From the results, all the strains were identified as P. yezoensis f. narawaensis, and P. tenera was not detected. Hence, it was confirmed that most of the Porphyra strains currently cultivated in Japan are P. yezoensis f. narawaensis, and that intensive selective breeding has led to a reduced genetic diversity in the stock used for nori cultivation. [ABSTRACT FROM AUTHOR]

Polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) analysis of the plastid ribulose-1,5-bisphosphate carboxylase (RuBisCo) spacer region was developed for a more reliable and rapid species identification of cultivated Porphyra in combination with PCR-RFLP analysis of the nuclear internal transcribed spacer (ITS) region. From the PCR-RFLP analyses of the plastid and nuclear DNA, we examined seven strains of conchocelis that were used for cultivation as Porphyra tenera Kjellman but without strict species identification. The PCR-RFLP analyses suggested that two strains, C-32 and 90-02, were cultivated P. tenera and that the other five strains, C-24, C-28, C-29, C-30 and M-1, were Porphyra yezoensis f. narawaensis Miura. To identify species more accurately and to reveal additional genetic variation, the two strains C-32 and 90-02 were further studied by sequencing their RuBisCo spacer and ITS-1 regions. Although RuBisCo spacer sequences of the two strains were identical to each other, each of their ITS-1 sequences showed a single substitution. The sequence data again confirmed that the two strains (C-32 and 90-02) were cultivated P. tenera, and suggested that the two strains showed some genetic variation. We concluded that PCR-RFLP analysis of the plastid and nuclear DNA is a powerful tool for reliable and rapid species identification of many strains of cultivated Porphyra in Japan and for the collection of genetically variable breeding material of Porphyra. [ABSTRACT FROM AUTHOR]

Porphyra teneraKjellman, widely cultivated in nori farms before the development of artificial seeding, is currently listed as an endangered species in Japan. To confirm whether a wild-collected gametophytic blade wasP. teneraor the closely related speciesP. yezoensisUeda, morphological observations and molecular analyses were made on the pure line HGT-1 isolated from a wild blade. This pure line was identified asP. tenerabased on detailed morphological features. Sequences of the nuclear internal transcribed spacer region 1 and the plastid RUBISCO spacer revealed thatP. teneraHGT-1 was clearly different fromP. yezoensisf.narawaensisMiura, the main species cultivated in Japan. PCR-RFLP analysis of the internal transcribed spacer region was found to be a convenient method for rapid discrimination betweenP. teneraand cultivatedP. yezoensis. The restriction patterns generated by the endonucleasesDraI andHaeIII were useful for differentiating between both gametophytic and conchocelis stages ofP. teneraHGT-1 andP. yezoensisf.narawaensisstrains. Thus, PCR-RFLP analysis will serve as a valuable tool for rapid species identification of cultivatedPorphyrastrains, culture collections ofPorphyrastrains for breeding material and conservation of biodiversity, and, as codominant cleaved amplified polymorphic sequence markers for interspecific hybridization products betweenP. teneraandP. yezoensisf.narawaensis. Under the same culture conditions, rate of blade length increase and the blade length-to-width ratio were lower inP. teneraHGT-1 than inP. yezoensisf.narawaensisHG-4. The HGT-1 became mature more rapidly than HG-4 and had thinner blades. [ABSTRACT FROM AUTHOR]

Detailed morphological observations were made on two strains of cultivated Porphyra: HG-1 (pure line isolated from Dai-1) and Noriken-4 (parental strain of a pure line HG-4) . The two strains were identified as P. yezoensis f. narawaensis based on their macroscopic and microscopic features, such as long linear or oblanceolate blades up to 50 cm in maximum length, division formulae of spermatangia and zygotosporangia, shape of trichogynes and carpogonia, and the second transverse divisional plane formed at the division from c/2 to c/4 in zygotosporangia. Gametophytic blades from two completely homozygous conchocelis strains isolated in this study (HG-1 and HG-4) were cultured under the same conditions and compared to confirm whether the differences in their shapes are genetically determined. The shape of blades from both of concho-spores and monospores was always more slender in HG-4 than in HG-1 at the same blade age, suggesting that the difference in the blade shape between the two pure lines is due to genetic variation. To estimate the level of genetic variation the two pure lines were subjected to amplified fragment length polymorphism fingerprint analysis. A total of 230 bands were detected in HG-1 and HG-4 using eight selective primer pairs, and the number of polymorphic bands was only two in HG-1. These results indicate that the two pure lines certainly show genetic variation, which is, however, at an extremely low level. The importance of pure-line breeding and the origin of currently cultivated Porphyra are discussed. This is the first report to identify currently cultivated Porphyra strains in Japan based on combined results of detailed morphological observations and molecular analysis. [ABSTRACT FROM AUTHOR]

In order to extract DNA rapidly from cultivated Porphyra, we extracted total DNA from conchocelis using the ISOPLANT II kit (Nippon Gene) without liquid nitrogen treatment or CsCl-gradient ultracentrifugation. By confirming the reproducibility of RAPD patterns, it is concluded that the quality of the extracted DNA is sufficient to use as a template for molecular investigation. Using this rapid method, the nuclear ribosomal DNA of the internal transcribed spacer (ITS) regions was amplified from seven strains of cultivated Porphyra, which had been maintained as free-living conchocelis by subculturing in the laboratory. From the amplified DNAs, the ITS-1 sequences were determined in order to identify the species and genetic relationship of the strains. The sequences were identical in the seven strains, and all the strains were identified as P. yezoensis. Furthermore, the gametophytic blades of these strains showed long linear or oblanceolate shapes in the laboratory culture. It was concluded that these strains are P. yezoensis form. narawaensis. This rapid DNA extraction method from conchocelis will be a powerful tool for phylogenetic analysis and for genetic improvement of cultivated Porphyra. [ABSTRACT FROM AUTHOR]

In order to characterize the spontaneous green-type pigmentation mutant of Porphyra yezoensis , growth and contents of photosynthetic pigments were compared with those of the wild type in gametophytic blades. The growth of the green mutant was slower than that of the wild type. The content of phycoerythrin was markedly lower in the green mutant than in the wild type. For genetic analysis, the green mutant was crossed with the wild type. In the cross, the heterozygous conchocelis (the wild-type color) produced many sectored F 1 gametophytic blades, which were composed of both parental colors. The segregation ratio by counting the wild-type sectors and the green-type sectors of the F 1 blades was approximately 1 : 1. This suggests that the green mutant is governed by a single recessive gene. All the monospore germlings from the sectored F 1 gametophytic blades that originated from the heterozygous conchocelis developed into single-colored blades with either the green color or the wild-type color. The advantages of using heterozygous conchocelis (hybrid conchocelis) and monospores from F 1 gametophytic blades that originated from the heterozygous conchocelis in commercial farming of Porphyra yezoensis are detailed and discussed. [ABSTRACT FROM AUTHOR]