Various in vitro grown tissues (non-regenerative callus, regenerative callus and microshoot deriv... more Various in vitro grown tissues (non-regenerative callus, regenerative callus and microshoot derived leaves) of Solanum nigrum L. were cultured under salinity stress (0-150 mM NaCl) for enhanced production of solasodine, a steroidal alkaloid and an alternative to diosgenin, which is used as a precursor for the commercial production of steroidal drugs. The role of plant growth regulators and various concentrations of NaCl during in vitro production of solasodine was studied. The in vitro yield was compared with the yield from leaves of field grown plant. Solasodine content was maximum (2.39 mg/g dry wt.) in regenerative callus when grown on medium added with 150 mM NaCl; followed by in vitro raised leaf of microshoot. Quantitative estimation of solasodine was carried out using a new HPTLC method, which is validated for its recovery and precession. The proposed HPTLC method showed a good linear relationship (r(2)=0.994) in 50-2000 ng/spot concentration ranges. The data demonstrate that the solasodine production in cultures was growth dependent.
Various in vitro grown tissues (non-regenerative callus, regenerative callus and microshoot deriv... more Various in vitro grown tissues (non-regenerative callus, regenerative callus and microshoot derived leaves) of Solanum nigrum L. were cultured under salinity stress (0-150 mM NaCl) for enhanced production of solasodine, a steroidal alkaloid and an alternative to diosgenin, which is used as a precursor for the commercial production of steroidal drugs. The role of plant growth regulators and various concentrations of NaCl during in vitro production of solasodine was studied. The in vitro yield was compared with the yield from leaves of field grown plant. Solasodine content was maximum (2.39 mg/g dry wt.) in regenerative callus when grown on medium added with 150 mM NaCl; followed by in vitro raised leaf of microshoot. Quantitative estimation of solasodine was carried out using a new HPTLC method, which is validated for its recovery and precession. The proposed HPTLC method showed a good linear relationship (r(2)=0.994) in 50-2000 ng/spot concentration ranges. The data demonstrate that the solasodine production in cultures was growth dependent.
Catharanthus roseus (L.) G. Don. (Apocynaceae) is an important dicotyledonous medicinal plant. It... more Catharanthus roseus (L.) G. Don. (Apocynaceae) is an important dicotyledonous medicinal plant. It produces vinblastine and vincristine, two alkaloids that are being used against a variety of cancers. In the present study, the freezing (−196, 4, 15°C) and non-freezing (25°C) temperature was imposed on embryogenic cultures, and later in vitro embryogeny and vinblastine production in C. roseus was studied. Somatic embryo (SE) production was maximum at 15°C, but the SE maturation was high at 4°C. The SEs, grown at 25°C, showed highest germination and plantlet conversion. Quantitative estimation of vinblastine was carried out using high-performance liquid chromatography in various in vitro raised tissues (embryogenic callus), embryo stages (proliferated, matured and germinated embryos)], and SE-derived plantlets (leaf, shoot, root and whole plant) after various freezing- and non-freezing temperature treatments. Vinblastine synthesis was temperature dependent in C. roseus that has been discussed in this present article.
Catharanthus roseus (L.) G. Don. (Apocynaceae) is an important dicotyledonous medicinal plant. It... more Catharanthus roseus (L.) G. Don. (Apocynaceae) is an important dicotyledonous medicinal plant. It produces vinblastine and vincristine, two alkaloids that are being used against a variety of cancers. In the present study, the freezing (−196, 4, 15°C) and non-freezing (25°C) temperature was imposed on embryogenic cultures, and later in vitro embryogeny and vinblastine production in C. roseus was studied. Somatic embryo (SE) production was maximum at 15°C, but the SE maturation was high at 4°C. The SEs, grown at 25°C, showed highest germination and plantlet conversion. Quantitative estimation of vinblastine was carried out using high-performance liquid chromatography in various in vitro raised tissues (embryogenic callus), embryo stages (proliferated, matured and germinated embryos)], and SE-derived plantlets (leaf, shoot, root and whole plant) after various freezing- and non-freezing temperature treatments. Vinblastine synthesis was temperature dependent in C. roseus that has been discussed in this present article.
Genotoxic studies with reference to pollution biomonitoring is a recent approach and can be used ... more Genotoxic studies with reference to pollution biomonitoring is a recent approach and can be used for maintenance of crop productivity. Effect of the cadmium chloride in the pure germ line of broad bean (Vicia faba L.) were evaluated in relation to the chromosomal abnormalities and rate of cell division. Seeds grown in the nutrient medium for 48 hrs containing different concentrations of cadmium chloride (control, 0.05 mM, 0.1 mM, 0.2 mM, 0.4 mM and 0.8 mM) showed different genotoxic effects such as polyploidy, multipolarity, chromosomal bridge with fragments, lagging chromosome and micronuclei. Relative division rate (RDR) was decreased with increasing cadmium concentration while relative abnormality rate (RAR) increased suggesting that cadmium may cause genetic changes in pure germ line and produce irrepairable damage in root meristem. Therefore, combining plant bioassays and root analysis of Cd++ content would allow us to define phytotoxicity thresholds in plants.
Genotoxic studies with reference to pollution biomonitoring is a recent approach and can be used ... more Genotoxic studies with reference to pollution biomonitoring is a recent approach and can be used for maintenance of crop productivity. Effect of the cadmium chloride in the pure germ line of broad bean (Vicia faba L.) were evaluated in relation to the chromosomal abnormalities and rate of cell division. Seeds grown in the nutrient medium for 48 hrs containing different concentrations of cadmium chloride (control, 0.05 mM, 0.1 mM, 0.2 mM, 0.4 mM and 0.8 mM) showed different genotoxic effects such as polyploidy, multipolarity, chromosomal bridge with fragments, lagging chromosome and micronuclei. Relative division rate (RDR) was decreased with increasing cadmium concentration while relative abnormality rate (RAR) increased suggesting that cadmium may cause genetic changes in pure germ line and produce irrepairable damage in root meristem. Therefore, combining plant bioassays and root analysis of Cd++ content would allow us to define phytotoxicity thresholds in plants.
In the present study an efficient somatic embryogenesis method has been developed in Catharanthus... more In the present study an efficient somatic embryogenesis method has been developed in Catharanthus roseus. Friable embryogenic callus was induced from hypocotyl of in vitro germinated seeds on Murashige and Skoog basal nutrient media supplemented with various auxins particularly 2,4-D (1.0 mg l−1). However, only NAA (1.0 mg l−1) produced somatic embryos in cultures. Embryo proliferation was even high on the same medium added with BAP. Cotyledonary somatic embryo germinated and converted into plantlets in BAP (0.5 mg l−1) added medium following a treatment with gibberellic acid (1.0 mg l−1) for maturation. Carbon sources and concentrations had a marked influence on maturation process. Plantlet conversion was better achieved when embryos were matured on 3% fructose or 3–6% maltose. The result discussed in this paper indicates that somatic embryos were produced in numbers and converted plantlets can be used as raw material, genetic modification to embryo precursor cell may improve alkaloid yield further.
Various in vitro grown tissues (non-regenerative callus, regenerative callus and microshoot deriv... more Various in vitro grown tissues (non-regenerative callus, regenerative callus and microshoot derived leaves) of Solanum nigrum L. were cultured under salinity stress (0-150 mM NaCl) for enhanced production of solasodine, a steroidal alkaloid and an alternative to diosgenin, which is used as a precursor for the commercial production of steroidal drugs. The role of plant growth regulators and various concentrations of NaCl during in vitro production of solasodine was studied. The in vitro yield was compared with the yield from leaves of field grown plant. Solasodine content was maximum (2.39 mg/g dry wt.) in regenerative callus when grown on medium added with 150 mM NaCl; followed by in vitro raised leaf of microshoot. Quantitative estimation of solasodine was carried out using a new HPTLC method, which is validated for its recovery and precession. The proposed HPTLC method showed a good linear relationship (r(2)=0.994) in 50-2000 ng/spot concentration ranges. The data demonstrate that the solasodine production in cultures was growth dependent.
In the present study an efficient somatic embryogenesis method has been developed in Catharanthus... more In the present study an efficient somatic embryogenesis method has been developed in Catharanthus roseus. Friable embryogenic callus was induced from hypocotyl of in vitro germinated seeds on Murashige and Skoog basal nutrient media supplemented with various auxins particularly 2,4-D (1.0 mg l−1). However, only NAA (1.0 mg l−1) produced somatic embryos in cultures. Embryo proliferation was even high on the same medium added with BAP. Cotyledonary somatic embryo germinated and converted into plantlets in BAP (0.5 mg l−1) added medium following a treatment with gibberellic acid (1.0 mg l−1) for maturation. Carbon sources and concentrations had a marked influence on maturation process. Plantlet conversion was better achieved when embryos were matured on 3% fructose or 3–6% maltose. The result discussed in this paper indicates that somatic embryos were produced in numbers and converted plantlets can be used as raw material, genetic modification to embryo precursor cell may improve alkaloid yield further.
In the present study, the regeneration pathway, especially the different events of somatic embryo... more In the present study, the regeneration pathway, especially the different events of somatic embryogenesis (SE) have been studied morphologically and biochemically in Catharanthus roseus. Firstly, the calluses were induced from different explant sources (hypocotyl, epicotyl and root) by using various auxins. Embryogenic and non-embryogenic calluses were identified based on their morphology, colour and dry weight. Embryogenic callus was later cultivated on MS added with 0.45 μM 2,4-D, 6.62 μM BAP and 1.44 μM GA3 for obtaining various developmental stages of embryos. Different stages of embryos have been assayed for the establishment of marker based embryogenesis, particularly on embryo specific proteins whose presence or absence will ensure a rapid and efficient production of embryos that has a special application to clonal biotechnology. Two embryo specific proteins (38 and 33 kD) have been identified for the first time in C. roseus during torpedo stage of embryogenesis. Besides, multiple shoot formation from in vitro raised emblings was also attempted to examine the role of BAP and kinetin for shoot proliferation. The shoots were rooted with 5.37 μM NAA and 5.71 μM IAA before transplantation.
In the present study, the regeneration pathway, especially the different events of somatic embryo... more In the present study, the regeneration pathway, especially the different events of somatic embryogenesis (SE) have been studied morphologically and biochemically in Catharanthus roseus. Firstly, the calluses were induced from different explant sources (hypocotyl, epicotyl and root) by using various auxins. Embryogenic and non-embryogenic calluses were identified based on their morphology, colour and dry weight. Embryogenic callus was later cultivated on MS added with 0.45 μM 2,4-D, 6.62 μM BAP and 1.44 μM GA3 for obtaining various developmental stages of embryos. Different stages of embryos have been assayed for the establishment of marker based embryogenesis, particularly on embryo specific proteins whose presence or absence will ensure a rapid and efficient production of embryos that has a special application to clonal biotechnology. Two embryo specific proteins (38 and 33 kD) have been identified for the first time in C. roseus during torpedo stage of embryogenesis. Besides, multiple shoot formation from in vitro raised emblings was also attempted to examine the role of BAP and kinetin for shoot proliferation. The shoots were rooted with 5.37 μM NAA and 5.71 μM IAA before transplantation.
Catharanthus roseus (L.) G. Don. (Apocynaceae) is an important dicotyledonous medicinal plant. It... more Catharanthus roseus (L.) G. Don. (Apocynaceae) is an important dicotyledonous medicinal plant. It produces vinblastine and vincristine, two alkaloids that are being used against a variety of cancers. In the present study, the freezing (−196, 4, 15°C) and non-freezing (25°C) temperature was imposed on embryogenic cultures, and later in vitro embryogeny and vinblastine production in C. roseus was studied. Somatic embryo (SE) production was maximum at 15°C, but the SE maturation was high at 4°C. The SEs, grown at 25°C, showed highest germination and plantlet conversion. Quantitative estimation of vinblastine was carried out using high-performance liquid chromatography in various in vitro raised tissues (embryogenic callus), embryo stages (proliferated, matured and germinated embryos)], and SE-derived plantlets (leaf, shoot, root and whole plant) after various freezing- and non-freezing temperature treatments. Vinblastine synthesis was temperature dependent in C. roseus that has been discussed in this present article.
Somatic embryos were induced from in vitro germinating seed-hypocotyls of Catharanthus roseus. Th... more Somatic embryos were induced from in vitro germinating seed-hypocotyls of Catharanthus roseus. The process of embryogenesis has been categorized into a few distinct stages (induction, proliferation, maturation and germination) in which liquid overlaying at varying levels 0 ml (T0), 0.25 ml (T1), 0.5 ml (T2), 0.75 ml (T3) and 1.0 ml (T4) was applied on solid medium. It was found that liquid overlaying improved proliferation; maturation, germination of embryos in C. roseus. In proliferation stage, particularly in T2, torpedo embryo number increased significantly (i.e. 129.6%) as compared to control. Liquid overlaying (T2, T3 and T4) also improved embryo maturation and showed early germination even in maturation medium. It also accelerated normal embryo germination frequency particularly in treatment with T2 and shortened ‘embryo—plantlet’ recovery time. Biochemical analyses revealed more proline, protein and amino acid with increasing level of liquid overlaying as it improved embryo induction, development and faster germination.
Somatic embryos were induced from in vitro germinating seed-hypocotyls of Catharanthus roseus. Th... more Somatic embryos were induced from in vitro germinating seed-hypocotyls of Catharanthus roseus. The process of embryogenesis has been categorized into a few distinct stages (induction, proliferation, maturation and germination) in which liquid overlaying at varying levels 0 ml (T0), 0.25 ml (T1), 0.5 ml (T2), 0.75 ml (T3) and 1.0 ml (T4) was applied on solid medium. It was found that liquid overlaying improved proliferation; maturation, germination of embryos in C. roseus. In proliferation stage, particularly in T2, torpedo embryo number increased significantly (i.e. 129.6%) as compared to control. Liquid overlaying (T2, T3 and T4) also improved embryo maturation and showed early germination even in maturation medium. It also accelerated normal embryo germination frequency particularly in treatment with T2 and shortened ‘embryo—plantlet’ recovery time. Biochemical analyses revealed more proline, protein and amino acid with increasing level of liquid overlaying as it improved embryo induction, development and faster germination.
Genotoxic studies with reference to pollution biomonitoring is a recent approach and can be used ... more Genotoxic studies with reference to pollution biomonitoring is a recent approach and can be used for maintenance of crop productivity. Effect of the cadmium chloride in the pure germ line of broad bean (Vicia faba L.) were evaluated in relation to the chromosomal abnormalities and rate of cell division. Seeds grown in the nutrient medium for 48 hrs containing different concentrations of cadmium chloride (control, 0.05 mM, 0.1 mM, 0.2 mM, 0.4 mM and 0.8 mM) showed different genotoxic effects such as polyploidy, multipolarity, chromosomal bridge with fragments, lagging chromosome and micronuclei. Relative division rate (RDR) was decreased with increasing cadmium concentration while relative abnormality rate (RAR) increased suggesting that cadmium may cause genetic changes in pure germ line and produce irrepairable damage in root meristem. Therefore, combining plant bioassays and root analysis of Cd++ content would allow us to define phytotoxicity thresholds in plants.
We established an efficient plant regeneration system for Catharanthus roseus L. (G.) Don through... more We established an efficient plant regeneration system for Catharanthus roseus L. (G.) Don through somatic embryogenesis. Embryogenic callus was induced from hypocotyl of seed germinated in vitro. Somatic embryogenesis in Catharanthus has been categorized into three distinct stages: (1) initiation and proliferation of embryo; (2) maturation, and; (3) germination or plantlet conversion. Beside plant growth regulators, various stages of embryogenesis were screened for their response to a wide variety of factors (pH, gelrite, light, sugar alcohols, polyethyleneglycol and amino acids), which affect embryogenesis. All of the tested factors had a small to marked influence on embryogeny and eventual conversion to plantlets. The plantlets were acclimatized successfully in a greenhouse. To our knowledge, this is the first report describing a detailed study of various cultural factors which regulate embryogenesis in C. roseus. The results discussed in this paper may be used in mass propagation to produce medicinal raw material, and the embryo precursor cells could be used in genetic modification programmes that aim to improve the alkaloid yield as well.
We established an efficient plant regeneration system for Catharanthus roseus L. (G.) Don through... more We established an efficient plant regeneration system for Catharanthus roseus L. (G.) Don through somatic embryogenesis. Embryogenic callus was induced from hypocotyl of seed germinated in vitro. Somatic embryogenesis in Catharanthus has been categorized into three distinct stages: (1) initiation and proliferation of embryo; (2) maturation, and; (3) germination or plantlet conversion. Beside plant growth regulators, various stages of embryogenesis were screened for their response to a wide variety of factors (pH, gelrite, light, sugar alcohols, polyethyleneglycol and amino acids), which affect embryogenesis. All of the tested factors had a small to marked influence on embryogeny and eventual conversion to plantlets. The plantlets were acclimatized successfully in a greenhouse. To our knowledge, this is the first report describing a detailed study of various cultural factors which regulate embryogenesis in C. roseus. The results discussed in this paper may be used in mass propagation to produce medicinal raw material, and the embryo precursor cells could be used in genetic modification programmes that aim to improve the alkaloid yield as well.
Various in vitro grown tissues (non-regenerative callus, regenerative callus and microshoot deriv... more Various in vitro grown tissues (non-regenerative callus, regenerative callus and microshoot derived leaves) of Solanum nigrum L. were cultured under salinity stress (0-150 mM NaCl) for enhanced production of solasodine, a steroidal alkaloid and an alternative to diosgenin, which is used as a precursor for the commercial production of steroidal drugs. The role of plant growth regulators and various concentrations of NaCl during in vitro production of solasodine was studied. The in vitro yield was compared with the yield from leaves of field grown plant. Solasodine content was maximum (2.39 mg/g dry wt.) in regenerative callus when grown on medium added with 150 mM NaCl; followed by in vitro raised leaf of microshoot. Quantitative estimation of solasodine was carried out using a new HPTLC method, which is validated for its recovery and precession. The proposed HPTLC method showed a good linear relationship (r(2)=0.994) in 50-2000 ng/spot concentration ranges. The data demonstrate that the solasodine production in cultures was growth dependent.
Various in vitro grown tissues (non-regenerative callus, regenerative callus and microshoot deriv... more Various in vitro grown tissues (non-regenerative callus, regenerative callus and microshoot derived leaves) of Solanum nigrum L. were cultured under salinity stress (0-150 mM NaCl) for enhanced production of solasodine, a steroidal alkaloid and an alternative to diosgenin, which is used as a precursor for the commercial production of steroidal drugs. The role of plant growth regulators and various concentrations of NaCl during in vitro production of solasodine was studied. The in vitro yield was compared with the yield from leaves of field grown plant. Solasodine content was maximum (2.39 mg/g dry wt.) in regenerative callus when grown on medium added with 150 mM NaCl; followed by in vitro raised leaf of microshoot. Quantitative estimation of solasodine was carried out using a new HPTLC method, which is validated for its recovery and precession. The proposed HPTLC method showed a good linear relationship (r(2)=0.994) in 50-2000 ng/spot concentration ranges. The data demonstrate that the solasodine production in cultures was growth dependent.
Catharanthus roseus (L.) G. Don. (Apocynaceae) is an important dicotyledonous medicinal plant. It... more Catharanthus roseus (L.) G. Don. (Apocynaceae) is an important dicotyledonous medicinal plant. It produces vinblastine and vincristine, two alkaloids that are being used against a variety of cancers. In the present study, the freezing (−196, 4, 15°C) and non-freezing (25°C) temperature was imposed on embryogenic cultures, and later in vitro embryogeny and vinblastine production in C. roseus was studied. Somatic embryo (SE) production was maximum at 15°C, but the SE maturation was high at 4°C. The SEs, grown at 25°C, showed highest germination and plantlet conversion. Quantitative estimation of vinblastine was carried out using high-performance liquid chromatography in various in vitro raised tissues (embryogenic callus), embryo stages (proliferated, matured and germinated embryos)], and SE-derived plantlets (leaf, shoot, root and whole plant) after various freezing- and non-freezing temperature treatments. Vinblastine synthesis was temperature dependent in C. roseus that has been discussed in this present article.
Catharanthus roseus (L.) G. Don. (Apocynaceae) is an important dicotyledonous medicinal plant. It... more Catharanthus roseus (L.) G. Don. (Apocynaceae) is an important dicotyledonous medicinal plant. It produces vinblastine and vincristine, two alkaloids that are being used against a variety of cancers. In the present study, the freezing (−196, 4, 15°C) and non-freezing (25°C) temperature was imposed on embryogenic cultures, and later in vitro embryogeny and vinblastine production in C. roseus was studied. Somatic embryo (SE) production was maximum at 15°C, but the SE maturation was high at 4°C. The SEs, grown at 25°C, showed highest germination and plantlet conversion. Quantitative estimation of vinblastine was carried out using high-performance liquid chromatography in various in vitro raised tissues (embryogenic callus), embryo stages (proliferated, matured and germinated embryos)], and SE-derived plantlets (leaf, shoot, root and whole plant) after various freezing- and non-freezing temperature treatments. Vinblastine synthesis was temperature dependent in C. roseus that has been discussed in this present article.
Genotoxic studies with reference to pollution biomonitoring is a recent approach and can be used ... more Genotoxic studies with reference to pollution biomonitoring is a recent approach and can be used for maintenance of crop productivity. Effect of the cadmium chloride in the pure germ line of broad bean (Vicia faba L.) were evaluated in relation to the chromosomal abnormalities and rate of cell division. Seeds grown in the nutrient medium for 48 hrs containing different concentrations of cadmium chloride (control, 0.05 mM, 0.1 mM, 0.2 mM, 0.4 mM and 0.8 mM) showed different genotoxic effects such as polyploidy, multipolarity, chromosomal bridge with fragments, lagging chromosome and micronuclei. Relative division rate (RDR) was decreased with increasing cadmium concentration while relative abnormality rate (RAR) increased suggesting that cadmium may cause genetic changes in pure germ line and produce irrepairable damage in root meristem. Therefore, combining plant bioassays and root analysis of Cd++ content would allow us to define phytotoxicity thresholds in plants.
Genotoxic studies with reference to pollution biomonitoring is a recent approach and can be used ... more Genotoxic studies with reference to pollution biomonitoring is a recent approach and can be used for maintenance of crop productivity. Effect of the cadmium chloride in the pure germ line of broad bean (Vicia faba L.) were evaluated in relation to the chromosomal abnormalities and rate of cell division. Seeds grown in the nutrient medium for 48 hrs containing different concentrations of cadmium chloride (control, 0.05 mM, 0.1 mM, 0.2 mM, 0.4 mM and 0.8 mM) showed different genotoxic effects such as polyploidy, multipolarity, chromosomal bridge with fragments, lagging chromosome and micronuclei. Relative division rate (RDR) was decreased with increasing cadmium concentration while relative abnormality rate (RAR) increased suggesting that cadmium may cause genetic changes in pure germ line and produce irrepairable damage in root meristem. Therefore, combining plant bioassays and root analysis of Cd++ content would allow us to define phytotoxicity thresholds in plants.
In the present study an efficient somatic embryogenesis method has been developed in Catharanthus... more In the present study an efficient somatic embryogenesis method has been developed in Catharanthus roseus. Friable embryogenic callus was induced from hypocotyl of in vitro germinated seeds on Murashige and Skoog basal nutrient media supplemented with various auxins particularly 2,4-D (1.0 mg l−1). However, only NAA (1.0 mg l−1) produced somatic embryos in cultures. Embryo proliferation was even high on the same medium added with BAP. Cotyledonary somatic embryo germinated and converted into plantlets in BAP (0.5 mg l−1) added medium following a treatment with gibberellic acid (1.0 mg l−1) for maturation. Carbon sources and concentrations had a marked influence on maturation process. Plantlet conversion was better achieved when embryos were matured on 3% fructose or 3–6% maltose. The result discussed in this paper indicates that somatic embryos were produced in numbers and converted plantlets can be used as raw material, genetic modification to embryo precursor cell may improve alkaloid yield further.
Various in vitro grown tissues (non-regenerative callus, regenerative callus and microshoot deriv... more Various in vitro grown tissues (non-regenerative callus, regenerative callus and microshoot derived leaves) of Solanum nigrum L. were cultured under salinity stress (0-150 mM NaCl) for enhanced production of solasodine, a steroidal alkaloid and an alternative to diosgenin, which is used as a precursor for the commercial production of steroidal drugs. The role of plant growth regulators and various concentrations of NaCl during in vitro production of solasodine was studied. The in vitro yield was compared with the yield from leaves of field grown plant. Solasodine content was maximum (2.39 mg/g dry wt.) in regenerative callus when grown on medium added with 150 mM NaCl; followed by in vitro raised leaf of microshoot. Quantitative estimation of solasodine was carried out using a new HPTLC method, which is validated for its recovery and precession. The proposed HPTLC method showed a good linear relationship (r(2)=0.994) in 50-2000 ng/spot concentration ranges. The data demonstrate that the solasodine production in cultures was growth dependent.
In the present study an efficient somatic embryogenesis method has been developed in Catharanthus... more In the present study an efficient somatic embryogenesis method has been developed in Catharanthus roseus. Friable embryogenic callus was induced from hypocotyl of in vitro germinated seeds on Murashige and Skoog basal nutrient media supplemented with various auxins particularly 2,4-D (1.0 mg l−1). However, only NAA (1.0 mg l−1) produced somatic embryos in cultures. Embryo proliferation was even high on the same medium added with BAP. Cotyledonary somatic embryo germinated and converted into plantlets in BAP (0.5 mg l−1) added medium following a treatment with gibberellic acid (1.0 mg l−1) for maturation. Carbon sources and concentrations had a marked influence on maturation process. Plantlet conversion was better achieved when embryos were matured on 3% fructose or 3–6% maltose. The result discussed in this paper indicates that somatic embryos were produced in numbers and converted plantlets can be used as raw material, genetic modification to embryo precursor cell may improve alkaloid yield further.
In the present study, the regeneration pathway, especially the different events of somatic embryo... more In the present study, the regeneration pathway, especially the different events of somatic embryogenesis (SE) have been studied morphologically and biochemically in Catharanthus roseus. Firstly, the calluses were induced from different explant sources (hypocotyl, epicotyl and root) by using various auxins. Embryogenic and non-embryogenic calluses were identified based on their morphology, colour and dry weight. Embryogenic callus was later cultivated on MS added with 0.45 μM 2,4-D, 6.62 μM BAP and 1.44 μM GA3 for obtaining various developmental stages of embryos. Different stages of embryos have been assayed for the establishment of marker based embryogenesis, particularly on embryo specific proteins whose presence or absence will ensure a rapid and efficient production of embryos that has a special application to clonal biotechnology. Two embryo specific proteins (38 and 33 kD) have been identified for the first time in C. roseus during torpedo stage of embryogenesis. Besides, multiple shoot formation from in vitro raised emblings was also attempted to examine the role of BAP and kinetin for shoot proliferation. The shoots were rooted with 5.37 μM NAA and 5.71 μM IAA before transplantation.
In the present study, the regeneration pathway, especially the different events of somatic embryo... more In the present study, the regeneration pathway, especially the different events of somatic embryogenesis (SE) have been studied morphologically and biochemically in Catharanthus roseus. Firstly, the calluses were induced from different explant sources (hypocotyl, epicotyl and root) by using various auxins. Embryogenic and non-embryogenic calluses were identified based on their morphology, colour and dry weight. Embryogenic callus was later cultivated on MS added with 0.45 μM 2,4-D, 6.62 μM BAP and 1.44 μM GA3 for obtaining various developmental stages of embryos. Different stages of embryos have been assayed for the establishment of marker based embryogenesis, particularly on embryo specific proteins whose presence or absence will ensure a rapid and efficient production of embryos that has a special application to clonal biotechnology. Two embryo specific proteins (38 and 33 kD) have been identified for the first time in C. roseus during torpedo stage of embryogenesis. Besides, multiple shoot formation from in vitro raised emblings was also attempted to examine the role of BAP and kinetin for shoot proliferation. The shoots were rooted with 5.37 μM NAA and 5.71 μM IAA before transplantation.
Catharanthus roseus (L.) G. Don. (Apocynaceae) is an important dicotyledonous medicinal plant. It... more Catharanthus roseus (L.) G. Don. (Apocynaceae) is an important dicotyledonous medicinal plant. It produces vinblastine and vincristine, two alkaloids that are being used against a variety of cancers. In the present study, the freezing (−196, 4, 15°C) and non-freezing (25°C) temperature was imposed on embryogenic cultures, and later in vitro embryogeny and vinblastine production in C. roseus was studied. Somatic embryo (SE) production was maximum at 15°C, but the SE maturation was high at 4°C. The SEs, grown at 25°C, showed highest germination and plantlet conversion. Quantitative estimation of vinblastine was carried out using high-performance liquid chromatography in various in vitro raised tissues (embryogenic callus), embryo stages (proliferated, matured and germinated embryos)], and SE-derived plantlets (leaf, shoot, root and whole plant) after various freezing- and non-freezing temperature treatments. Vinblastine synthesis was temperature dependent in C. roseus that has been discussed in this present article.
Somatic embryos were induced from in vitro germinating seed-hypocotyls of Catharanthus roseus. Th... more Somatic embryos were induced from in vitro germinating seed-hypocotyls of Catharanthus roseus. The process of embryogenesis has been categorized into a few distinct stages (induction, proliferation, maturation and germination) in which liquid overlaying at varying levels 0 ml (T0), 0.25 ml (T1), 0.5 ml (T2), 0.75 ml (T3) and 1.0 ml (T4) was applied on solid medium. It was found that liquid overlaying improved proliferation; maturation, germination of embryos in C. roseus. In proliferation stage, particularly in T2, torpedo embryo number increased significantly (i.e. 129.6%) as compared to control. Liquid overlaying (T2, T3 and T4) also improved embryo maturation and showed early germination even in maturation medium. It also accelerated normal embryo germination frequency particularly in treatment with T2 and shortened ‘embryo—plantlet’ recovery time. Biochemical analyses revealed more proline, protein and amino acid with increasing level of liquid overlaying as it improved embryo induction, development and faster germination.
Somatic embryos were induced from in vitro germinating seed-hypocotyls of Catharanthus roseus. Th... more Somatic embryos were induced from in vitro germinating seed-hypocotyls of Catharanthus roseus. The process of embryogenesis has been categorized into a few distinct stages (induction, proliferation, maturation and germination) in which liquid overlaying at varying levels 0 ml (T0), 0.25 ml (T1), 0.5 ml (T2), 0.75 ml (T3) and 1.0 ml (T4) was applied on solid medium. It was found that liquid overlaying improved proliferation; maturation, germination of embryos in C. roseus. In proliferation stage, particularly in T2, torpedo embryo number increased significantly (i.e. 129.6%) as compared to control. Liquid overlaying (T2, T3 and T4) also improved embryo maturation and showed early germination even in maturation medium. It also accelerated normal embryo germination frequency particularly in treatment with T2 and shortened ‘embryo—plantlet’ recovery time. Biochemical analyses revealed more proline, protein and amino acid with increasing level of liquid overlaying as it improved embryo induction, development and faster germination.
Genotoxic studies with reference to pollution biomonitoring is a recent approach and can be used ... more Genotoxic studies with reference to pollution biomonitoring is a recent approach and can be used for maintenance of crop productivity. Effect of the cadmium chloride in the pure germ line of broad bean (Vicia faba L.) were evaluated in relation to the chromosomal abnormalities and rate of cell division. Seeds grown in the nutrient medium for 48 hrs containing different concentrations of cadmium chloride (control, 0.05 mM, 0.1 mM, 0.2 mM, 0.4 mM and 0.8 mM) showed different genotoxic effects such as polyploidy, multipolarity, chromosomal bridge with fragments, lagging chromosome and micronuclei. Relative division rate (RDR) was decreased with increasing cadmium concentration while relative abnormality rate (RAR) increased suggesting that cadmium may cause genetic changes in pure germ line and produce irrepairable damage in root meristem. Therefore, combining plant bioassays and root analysis of Cd++ content would allow us to define phytotoxicity thresholds in plants.
We established an efficient plant regeneration system for Catharanthus roseus L. (G.) Don through... more We established an efficient plant regeneration system for Catharanthus roseus L. (G.) Don through somatic embryogenesis. Embryogenic callus was induced from hypocotyl of seed germinated in vitro. Somatic embryogenesis in Catharanthus has been categorized into three distinct stages: (1) initiation and proliferation of embryo; (2) maturation, and; (3) germination or plantlet conversion. Beside plant growth regulators, various stages of embryogenesis were screened for their response to a wide variety of factors (pH, gelrite, light, sugar alcohols, polyethyleneglycol and amino acids), which affect embryogenesis. All of the tested factors had a small to marked influence on embryogeny and eventual conversion to plantlets. The plantlets were acclimatized successfully in a greenhouse. To our knowledge, this is the first report describing a detailed study of various cultural factors which regulate embryogenesis in C. roseus. The results discussed in this paper may be used in mass propagation to produce medicinal raw material, and the embryo precursor cells could be used in genetic modification programmes that aim to improve the alkaloid yield as well.
We established an efficient plant regeneration system for Catharanthus roseus L. (G.) Don through... more We established an efficient plant regeneration system for Catharanthus roseus L. (G.) Don through somatic embryogenesis. Embryogenic callus was induced from hypocotyl of seed germinated in vitro. Somatic embryogenesis in Catharanthus has been categorized into three distinct stages: (1) initiation and proliferation of embryo; (2) maturation, and; (3) germination or plantlet conversion. Beside plant growth regulators, various stages of embryogenesis were screened for their response to a wide variety of factors (pH, gelrite, light, sugar alcohols, polyethyleneglycol and amino acids), which affect embryogenesis. All of the tested factors had a small to marked influence on embryogeny and eventual conversion to plantlets. The plantlets were acclimatized successfully in a greenhouse. To our knowledge, this is the first report describing a detailed study of various cultural factors which regulate embryogenesis in C. roseus. The results discussed in this paper may be used in mass propagation to produce medicinal raw material, and the embryo precursor cells could be used in genetic modification programmes that aim to improve the alkaloid yield as well.
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