This investigation reports a protocol for transfer and expression of foreign chimeric genes in loblolly pine (Pinus taeda L.). Transformation was achieved by co-cultivation of mature zygotic embryos with Agrobacterium...This investigation reports a protocol for transfer and expression of foreign chimeric genes in loblolly pine (Pinus taeda L.). Transformation was achieved by co-cultivation of mature zygotic embryos with Agrobacterium tumefaciens strain LBA4404 which harbored a binary vector (pBI121) including genes for β-glucuronidase (GUS) and neomycin phosphotransferase (NPTII). Factors influencing transgene expression including seed sources of loblolly pine, concentration of bacteria, and the wounding procedures of target explants were investigated. The expression of foreign gene was confirmed by the ability of mature zygotic embryos to produce calli in the presence of kanamycin, by histochemical assays of GUS activity, by PCR analysis, and by Southern blot. The successful expression of the GUS gene in different families of loblolly pine suggests that this transformation system is probably useful for the production of the genetically modified conifers.展开更多
A protocol is presented for genetically engineering loblolly pine (Pinus taeda L.) using particle bombardment. This protocol enabled the routine transformation of loblolly pine plants that were previously difficult to...A protocol is presented for genetically engineering loblolly pine (Pinus taeda L.) using particle bombardment. This protocol enabled the routine transformation of loblolly pine plants that were previously difficult to transform. Mature zygotic embryos were used to be bombarded and to generate organogenic callus and transgenic regenerated plants. Plasmid pB48.215 DNA contained a synthetic Bacillus thuringiensis (B.t.) cryIAc coding sequence flanked by the double cauliflower mosaic virus (CaMV) 35S promoter and nopaline synthase (Nos) terminator sequences, and the selectable marker gene, neomycin phosphotransferase II (nptII) controlled by the promoter of the nopaline synthase gene was introduced into loblolly pine tissues by particle bombardment. The transformed tissues were proliferated and selected by kanamycin resistance conferred by the introduced NPTII gene. Shoot regeneration was induced from the kanamycin-resistant callus, and transgenic plantlets were then produced. The presence of the introduced genes in the transgenic loblolly pine plants was confirmed by polymerase chain reactions (PCR) analysis, by Southern blot analysis, and insect feeding assays. The recovered transgenic plants were acclimatized and then established in soil.展开更多
Three antibiotics ampicillin, carbenicillin, and cefotaxime were evaluated for their effects on induction, growth, and differentiation of organogenic calli, as well as rooting of regenerated shoots of three loblolly p...Three antibiotics ampicillin, carbenicillin, and cefotaxime were evaluated for their effects on induction, growth, and differentiation of organogenic calli, as well as rooting of regenerated shoots of three loblolly pine (Pinus taeda L.) genotypes. Of the antibiotics administered, cefotaxime maximally increased the frequency of callus formation and growth rate of organogenic calli, carbenicillin maximally increased the frequency of shoot regeneration and the average number of adventitious shoots per piece of organogenic callus, ampicillin maximally decreased the rooting frequency of regenerated shoots and mean number of roots per regenerated shoot, in comparison with antibiotic-free media. Compared with the control, ampicillin minimally increased the frequency of callus formation, cefotaxime minimally increased the frequency of shoot regeneration, and carbenicillin mini-mally decreased the rooting frequency of regenerated shoots in three loblolly pine genotypes tested. All three antibiotics in-creased the frequencies of callus formation and shoot regeneration, and reduced the rooting frequency of regenerated shoots suggested that the establishment of an efficient Agrobacterium tumefaciens-mediated transformation protocol for stable integra-tion of foreign genes into loblolly pine need to select a suitable antibiotic. This investigation could be useful for optimizing genetic transformation of conifers.展开更多
Protoplasts of embryogenic suspension cells of loblolly pine (Pinus taeda L).were isolated at exponential growth stage.Influences of various concentrations of basal medium,levels of BA,and concentrations of inositol ...Protoplasts of embryogenic suspension cells of loblolly pine (Pinus taeda L).were isolated at exponential growth stage.Influences of various concentrations of basal medium,levels of BA,and concentrations of inositol on the differentiation of embryonal suspensor mass (ESM),early stage somatic embryos (ESE) ,and lae stage somatic embryos (LSE) were investigated .A study of the effect of various concentrations of LP basal medium sowed that the optimal basal medium concentration of ESM,ESE,and LSE differentiation was 1.25 LP medium.The effects of various levels of BA and inositol showed that the optimal concentrations of BA for the formation of ESM,ESE and LSE were 4 mg/L ,2mg/L and 1mg/L,respectively ,and the optimal concentrations of inositol for the ESM ,ESE and LSM formation were 400mg/L,800mg/L and 1,200mg/L,respectively.展开更多
Trials of 16-26 years old progenies from Yingde Loblolly Pine Seed Orchard were investigated by sample plot investigation.The trial fields were located at Yingde Loblolly Pine Seed Orchard,Yingde Forest Research Insti...Trials of 16-26 years old progenies from Yingde Loblolly Pine Seed Orchard were investigated by sample plot investigation.The trial fields were located at Yingde Loblolly Pine Seed Orchard,Yingde Forest Research Institute,Jiuqushui Forest Farm of Wengyuan County,Shaoguan Forest Farm and Renhua Forest Research Institute respectively.The growth process and performance level were studied by stem analysis based on average trees.Compared with that of local Masson pine(Pinus massoniana),the seedlots of loblolly pine(Pinus taeda) from Hubei and the seedlot of slash pine(P.elliottii) from Taishan Slash Pine Seed Orchard,the growth potential of progenies from Yingde Loblolly Pine Seed Orchard in North Guangdong was comprehensively evaluated.The results are as follows: The fast-growing period of height for the progenies from Yingde Loblolly Pine Seed Orchard began at the age of 3 and lasted for five to nine years.For diameter at breast-height(DBH) it began at the age of 5 and lasted for five to eight years.The fast-growing period for volume in Yingde Loblolly Pine Seed Orchard began at the age of 10 and lasted to 25 years old,while it began at the age of 10 both in Yingde Forest Research Institute and Jiuqushui Forest Farm,and at the age of 12 both in Shaoguan Forest Farm and Renhua Forest Research Institute.The progenies from Yingde Loblolly Pine Seed Orchard performed well generally in North Guangdong.Shaoguan and Renhua were suitable for the cultivation of large-dimension timber with the rotation longer than 25 years.In order to cultivate largedimension timber,suitable fertilizer shall be used when the DBH growth reaches the fast-growing period,and thinning operation shall be conducted when the mean annual increment for DBH reaches the maximum.展开更多
文摘This investigation reports a protocol for transfer and expression of foreign chimeric genes in loblolly pine (Pinus taeda L.). Transformation was achieved by co-cultivation of mature zygotic embryos with Agrobacterium tumefaciens strain LBA4404 which harbored a binary vector (pBI121) including genes for β-glucuronidase (GUS) and neomycin phosphotransferase (NPTII). Factors influencing transgene expression including seed sources of loblolly pine, concentration of bacteria, and the wounding procedures of target explants were investigated. The expression of foreign gene was confirmed by the ability of mature zygotic embryos to produce calli in the presence of kanamycin, by histochemical assays of GUS activity, by PCR analysis, and by Southern blot. The successful expression of the GUS gene in different families of loblolly pine suggests that this transformation system is probably useful for the production of the genetically modified conifers.
文摘A protocol is presented for genetically engineering loblolly pine (Pinus taeda L.) using particle bombardment. This protocol enabled the routine transformation of loblolly pine plants that were previously difficult to transform. Mature zygotic embryos were used to be bombarded and to generate organogenic callus and transgenic regenerated plants. Plasmid pB48.215 DNA contained a synthetic Bacillus thuringiensis (B.t.) cryIAc coding sequence flanked by the double cauliflower mosaic virus (CaMV) 35S promoter and nopaline synthase (Nos) terminator sequences, and the selectable marker gene, neomycin phosphotransferase II (nptII) controlled by the promoter of the nopaline synthase gene was introduced into loblolly pine tissues by particle bombardment. The transformed tissues were proliferated and selected by kanamycin resistance conferred by the introduced NPTII gene. Shoot regeneration was induced from the kanamycin-resistant callus, and transgenic plantlets were then produced. The presence of the introduced genes in the transgenic loblolly pine plants was confirmed by polymerase chain reactions (PCR) analysis, by Southern blot analysis, and insect feeding assays. The recovered transgenic plants were acclimatized and then established in soil.
文摘Three antibiotics ampicillin, carbenicillin, and cefotaxime were evaluated for their effects on induction, growth, and differentiation of organogenic calli, as well as rooting of regenerated shoots of three loblolly pine (Pinus taeda L.) genotypes. Of the antibiotics administered, cefotaxime maximally increased the frequency of callus formation and growth rate of organogenic calli, carbenicillin maximally increased the frequency of shoot regeneration and the average number of adventitious shoots per piece of organogenic callus, ampicillin maximally decreased the rooting frequency of regenerated shoots and mean number of roots per regenerated shoot, in comparison with antibiotic-free media. Compared with the control, ampicillin minimally increased the frequency of callus formation, cefotaxime minimally increased the frequency of shoot regeneration, and carbenicillin mini-mally decreased the rooting frequency of regenerated shoots in three loblolly pine genotypes tested. All three antibiotics in-creased the frequencies of callus formation and shoot regeneration, and reduced the rooting frequency of regenerated shoots suggested that the establishment of an efficient Agrobacterium tumefaciens-mediated transformation protocol for stable integra-tion of foreign genes into loblolly pine need to select a suitable antibiotic. This investigation could be useful for optimizing genetic transformation of conifers.
文摘Protoplasts of embryogenic suspension cells of loblolly pine (Pinus taeda L).were isolated at exponential growth stage.Influences of various concentrations of basal medium,levels of BA,and concentrations of inositol on the differentiation of embryonal suspensor mass (ESM),early stage somatic embryos (ESE) ,and lae stage somatic embryos (LSE) were investigated .A study of the effect of various concentrations of LP basal medium sowed that the optimal basal medium concentration of ESM,ESE,and LSE differentiation was 1.25 LP medium.The effects of various levels of BA and inositol showed that the optimal concentrations of BA for the formation of ESM,ESE and LSE were 4 mg/L ,2mg/L and 1mg/L,respectively ,and the optimal concentrations of inositol for the ESM ,ESE and LSM formation were 400mg/L,800mg/L and 1,200mg/L,respectively.
文摘Trials of 16-26 years old progenies from Yingde Loblolly Pine Seed Orchard were investigated by sample plot investigation.The trial fields were located at Yingde Loblolly Pine Seed Orchard,Yingde Forest Research Institute,Jiuqushui Forest Farm of Wengyuan County,Shaoguan Forest Farm and Renhua Forest Research Institute respectively.The growth process and performance level were studied by stem analysis based on average trees.Compared with that of local Masson pine(Pinus massoniana),the seedlots of loblolly pine(Pinus taeda) from Hubei and the seedlot of slash pine(P.elliottii) from Taishan Slash Pine Seed Orchard,the growth potential of progenies from Yingde Loblolly Pine Seed Orchard in North Guangdong was comprehensively evaluated.The results are as follows: The fast-growing period of height for the progenies from Yingde Loblolly Pine Seed Orchard began at the age of 3 and lasted for five to nine years.For diameter at breast-height(DBH) it began at the age of 5 and lasted for five to eight years.The fast-growing period for volume in Yingde Loblolly Pine Seed Orchard began at the age of 10 and lasted to 25 years old,while it began at the age of 10 both in Yingde Forest Research Institute and Jiuqushui Forest Farm,and at the age of 12 both in Shaoguan Forest Farm and Renhua Forest Research Institute.The progenies from Yingde Loblolly Pine Seed Orchard performed well generally in North Guangdong.Shaoguan and Renhua were suitable for the cultivation of large-dimension timber with the rotation longer than 25 years.In order to cultivate largedimension timber,suitable fertilizer shall be used when the DBH growth reaches the fast-growing period,and thinning operation shall be conducted when the mean annual increment for DBH reaches the maximum.
