Abstract
The interest in developing tissue culture-independent genetic transformation methods for plants has been growth. The pollen-tube pathway transformation technique is one method; however, this method is controversial because it is difficult to duplicate and produces insufficient molecular evidence to confirm transformation. Our objective was to evaluate the robustness of the soybean pollen-tube pathway technique (Glycine max L. Merr.). Solutions of purified DNA constructs carrying abar marker gene and agus reporter gene or a gene of interest (npk1) were applied to severed styles of flowers 6–8 h after self-pollination. The experiment was repeated 3 summers in the field, in which 4 DNA constructs and 7 soybean genotypes were tested. A total of 4793 progeny seeds were harvested from 5590 individually treated soybean flowers. All seeds were germinated and screened for transformants with herbicide spray, histochemical GUS assay, and Southern blot analysis. Although 2% of progenies showed partial resistance to the herbicide, no positive plants were identified from GUS assay and Southern analysis. Our results indicate that soybean pollen-tube pathway transformation is not reproducible.
Similar content being viewed by others
Abbreviations
- RAPD:
-
random amplified polymorphic DNA
References
Bent AF (2000) Arabidopsisin planta transformation: Uses, mechanisms, and prospects for transformation of other species. Plant Physiol 124: 1540–1547.
Birch RG (1997) Plant transformation: Problems and strategies for practical application. Annu Rev Plant Physiol. Plant Mol Biol 48: 297–326.
Carlson JB and Lersten NR (1987) Reproductive morphology. In: JR Wilcox (ed), Soybean: Improvement, production and uses. 2nd ed. Agronomy 16, pp 95–134.
Chen WS, Chiu CC, Liu HY, Lee TL, Chen JT, Lin CC, Wu YJ, and Chang HY (1998) Gene transfer via pollen-tube pathway for anti-fusarium wilt in watermelon. Biochem Mol Biol Int 46: 1201–1209.
Christou P (1995) Strategies for variety-independent genetic transformation of important cereals, legumes, and woody species utilizing particle bombardment. Euphytica 85: 13–27.
Duan XL and Chen SB (1985) Variation occurring by introduction of the exogenous DNA into rice. Sci Agric Sin 18: 6–10.
Frame B, Shou HX, Chikwamba R, Zhang ZY, Xiang CB, Fonger T, Pegg SE, Li BC, Nettleton D, Pei DQ, and Wang K (2002)Agrobacterium-mediated transformation of maize embryo using standard binary vector system. Plant Physiol 129: 13–22.
Gerlach WL and Bedbrook JR (1979) Cloning and characterization of ribosomal RNA genes from wheat and barley. Nucl Acid Res 7: 1869–1885.
Hu CY and Wang LZ (1999)In planta soybean transformation technologies developed in China: Procedure, confirmation and field performance. In Vitro Cell Dev Biol Plant 35: 417–420.
Jefferson RA, Kavanagh TA, and Bevan MW (1987) Gus fusion: β-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J 6: 3901–3907.
Kovtun Y, Chiu WL, Tena G, and Sheen J (2000) Functional analysis of oxidative stress-activated MAPK cascade in plants. Proc Natl Acad Sci USA. 97: 2940–2945.
Lei BJ, Li XC, Lu CH, Qian H, Zhou SJ, Xie WW, and Wang B (1994) Introduction of wild soybean DNA into cultivar soybean and molecular RAPD confirmation. Sci China (Ser B) 24: 596–601.
Lei BJ, Lu CH, Qian H, Li XC, Lu YB, Zhao K, Liu GY, and Yang XY (1995) New soy-bean strains of high protein and double high content obtained from introduction of exogenous total DNA. Soybean Sci 14: 203–208.
Liu DP, Yuan Y, and Sun H (1992) A study of exogenous DNA introduction into cultivated soybean. In: Zhou GY, Chen SB, and Hu JQ (eds), Advances in Molecular Breeding Research of Agriculture, pp 134–140, China Agri Sci Tech Press.
Liu DP, Liao L, Yuan Y, and Liu YZ (1997) Gaining of soybean lines resistant to SMV by introducing exogenous DNA. Soybean Sci 16: 277–281.
Luo ZX and Wu R (1989) A simple method for the transformation of rice via the pollentube pathway. Plant Mol Biol Rep 7: 69–77.
Moore S, Croughan T, Myers G and Vidrine R (1996) Investigation of transferring the BAR gene into soybean via the pollen tube pathway. Soybean Genet Newsl 23: 167–168.
Nagy F, Odell JT, Morelli G, and Chua NH (1985) Properties of expression of the 35S promoter from CaMV in transgenic tobacco plants. In: Zaitlin M, Day P, and Hollaender A (eds), Biotechnology in Plant Science: Relevance to Agriculture in the Eighties, pp 227–235. Academic Press, Orlando, FL.
Ni WC, Zhang ZL, and Guo SD (1998) Development of transgenic insect-resistant cotton plants. Sci Agric Sin 31: 8–13.
Potrykus I (1990) Gene transfer to cereals: An assessment. Bio/Technology 8: 535–542.
Reichardt M and Rogers S (1994) Preparation of plant DNA using CTAB. In: Ausubel F, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, and Struhl K (eds), Current Protocols in Molecular Biology, pp 2.3.3–2.3.7 John Wiley and Sons, New York.
Southern EM (1975) Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol 98: 503–517.
Widholm J (1999) Status of soybean transformation methods. In: Kauffman HE (ed), World Soybean Res Conf VI, pp 62–67, Superior Printing, Champaign, IL.
Yu HX, Liu JJ, Feng ZL, and Dong JD (1999) Study on introduction of vermin-resistance gene (CpTI) into wheat through pollen-tube pathway method. J ShanDong Agric Sci 99: 5–8.
Zhao LM, Liu DP, Sun H, Yun Y, and Huang M (1995) A sterile material of soybean gained by introducing exogenous DNA. Soybean Sci 14: 83–87.
Zhang Z, Aiqui X, Staswick P and Clemente TE (1999) The use of glufosinate as a selective agent inAgrobacterium-mediated transformation of soybean. Plant Cell Tissue Org Cult 56: 37–46.
Zhen JZ, Wu YX, Wang DJ, Zhang J, Ma ZR, and Zhou ZY (1998) The exploration of the mechanism and genetic performance of the progenies gained from pollen-tube pathway transformation. Sci Bull (China) 43: 561–566.
Zhou GY, Weng J, Zhen YS, Huang JG, Qian SY, and Liu GL (1983) Introduction of exogenous DNA into cotton embryos. In: Wu R, Grossman L and Molddave K (eds), Methods in enzymology, Vol 101, Recombination DNA, Part C, pp 433–481. Academic Press, New York.
Zhou GY, Weng J, Gong ZZ, Zhen YS, Yang WX, Shen WF, Wang ZF, Tao QZ, Huang JG, Qian SY, Liu GL, Ying MC, Xue DY, Hong AH, Xu YJ, Chen SB, and Duan XL (1988) Molecular breeding of agriculture: A technique for introducing exogenous DNA into plants after pollination. Sci Agric Sin 21: 1–6.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Shou, H., Palmer, R.G. & Wang, K. Irreproducibility of the soybean pollen-tube pathway transformation procedure. Plant Mol Biol Rep 20, 325–334 (2002). https://doi.org/10.1007/BF02772120
Published:
Issue Date:
DOI: https://doi.org/10.1007/BF02772120