Methods for tissue culturing and transforming elite inbreds of Zea mays L.

a technology of zea mays and tissue culture, which is applied in the field of tissue culture and can solve the problems of short time taken to produce transgenic plants and do not allow efficient transformation of elite lines, and achieve the effects of enhancing the transformation of elite corn inbreds, enhancing the integration of foreign dna into corn tissue, and high frequency

Inactive Publication Date: 2005-12-15
WILSON HERBERT M +1
View PDF13 Cites 4 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0012] In accordance with a second embodiment of the present invention, the integration of foreign DNA into corn tissue in transformation of corn tissue is enhanced by application of a heat shock treatment following contact of the foreign DNA with the corn tissue. In a preferred embodiment, the heat shock is conducted at a temperature of 45° C. for about 30 to about 180 minutes, preferably about 30 to about 60 minutes, more preferably about 30 minutes, at a time from about 24 to about 72 hours after the contact of the DNA with the corn tissue, preferably from 48 to 54 hours.
[0013] In a third embodiment of the present invention, the transformation of elite corn inbreds is achieved at a high frequency by co-cultivating corn tissue with Agrobacterium tumefaciens. In one particular embodiment, the transformation of elite corn inbreds is enhanced by using Agrobacterium freshly grown from glycerol cultures stored at about −86° C. In a second specific embodiment, the frequency of transformation is enhanced by co-cultivating corn tissue and Agrobacterium at 19° C. In a third particular embodiment, the frequency of transformation is enhanced by using lower levels of cefotaxime in the culturing media. In a further embodiment, the frequency of transformation of elite corn inbreds is enhanced using a combination of all of these techniques.

Problems solved by technology

Firstly, the time taken to produce transgenic plants is short when compared to other methods.
Initial methods of Agrobacterium-mediated corn transformation which were developed, while effective for some germplasm, do not allow for efficient transformation of elite lines.

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Examples

Experimental program
Comparison scheme
Effect test

example 1

Improvement in the Induction Frequency of Type II Callus

[0040] Plants of the elite corn inbred Stine 963 were grown under controlled conditions, either in a grow room or in a greenhouse. Plants were exposed throughout growth to a 16 h photoperiod with a daytime temperature of around 30° C. and a nighttime temperature of around 25° C. Plants were selfed and after approximately 10 days ears were harvested. At this time the immature embryos were on average between 1.0 and 2.0 mm in length. Usually the ears were then placed at 4° for two days prior to immature embryo excision.

[0041] The response on standard N6A media (Table 1) for inducing Type II callus was variable and appeared to be dependent on the physiological state of the donor plant. The frequency of Type II callus ranged from 0% up to 95%, with an average of around 40%. Immature embryos were excised with a view to using them as targets for Agrobacterium-mediated DNA delivery. For this work a reproducible and high frequency re...

example 2

Transformation of an Elite Corn Inbred by Agrobacterium tumefaciens

[0046] Modifications to the protocol of Hei and Komari (1997) involving co-cultivation temperature, culture media, antibiotic concentrations and Agrobacterium source cultures.

[0047]Agrobacterium strain LBA 4404 harboring “super binary” vectors as described in U.S. patent Hei and Komari (1997) was used in corn transformation experiments. Vectors with a bar expression cassette from pBARGUS (Fromm et. al., 1990) were used to generate resistance to the herbicide bialaphos, and a gus expression cassette from plG221 (Ohta et al., 1990) was used to produce Gus expression for transient assays. The gus expression cassette contains an intron in the N-terminal region of the gus gene which prevents expression in bacteria, but upon expression in plant cells the intron is spliced out and Gus activity is achieved (Ohta et al., 1990; Ishida et al., 1996). Agrobacterium containing “super binary” vectors were stored in glycerol stoc...

example 3

Effect of Heat Shock Treatment on Integration of DNA

[0053] Use of a brief heat treatment induces a transient state of so-called ‘competence’ in bacteria, allowing them to take up and express DNA from a variety of sources (cited in Sambrook et al, 1989). Use of a heat shock treatment to improve transformation efficiencies in higher organisms has not been reported. It was decided to explore the possibility that a heat shock treatment could improve integration of DNA following uptake. This was investigated with Agrobacterium-mediated DNA delivery in the first instance. First, a heat shock treatment of 45° for 30 minutes was administered to immature embryos 21, 24, 27, and 30 hours after the initiation of co-cultivation with Agrobacterium on LSAS (Table 1). No enhancement of clone production was noted after 21 hours, but promising preliminary results were obtained with the longer time periods. Further experiments were then performed—the results are presented in Table 6.

TABLE 6Effect ...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

PropertyMeasurementUnit
temperatureaaaaaaaaaa
temperatureaaaaaaaaaa
temperatureaaaaaaaaaa
Login to view more

Abstract

The present invention is directed to methods for the tissue culture and transformation of elite inbreds of corn (Zea mays L.). More specifically, the present invention is directed to a method for initiating Type II callus from corn tissue. The present invention is also directed to a method for enhancing the integration of foreign DNA in the transformation of corn using a heat shock treatment. The present invention is further directed to a method of transforming elite inbreds of corn using Agrobacterium.

Description

CROSS REFERENCE [0001] This application is a divisional of U.S. patent application having Ser. No. 09 / 203,679 filed Dec. 1, 1998 that has matured into U.S. Pat. No. 6,420,630 issued Jul. 16, 2002.BACKGROUND OF THE INVENTION [0002] The present invention is directed to methods for the tissue culture and transformation of elite inbreds of corn (Zea mays L.). More specifically, the present invention is directed to a method for initiating Type II callus from corn tissue. The present invention is also directed to a method for enhancing the integration of foreign DNA in the transformation of corn using a heat shock treatment. The present invention is further directed to a method of transforming elite inbreds of corn using Agrobacterium. [0003] The publications and other materials used herein to illuminate the background of the invention, and in particular, cases to provide additional details respecting the practice, are incorporated by reference, and for convenience are referenced in the f...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
Patent Type & Authority Applications(United States)
IPC IPC(8): A01H4/00C12N15/82
CPCC12N15/8205A01H4/008
Inventor WILSON, HERBERT M.HELD, BRUCE MARVIN
Owner WILSON HERBERT M
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap