Generation of doubled haploid transgenic wheat lines by microspore transformation
Brew-Appiah, RA; Ankrah, N; Liu, W; Konzak, CF; von Wettstein, D; Rustgi, S
HERO ID
3539426
Reference Type
Journal Article
Year
2013
Language
English
PMID
| HERO ID | 3539426 |
|---|---|
| In Press | No |
| Year | 2013 |
| Title | Generation of doubled haploid transgenic wheat lines by microspore transformation |
| Authors | Brew-Appiah, RA; Ankrah, N; Liu, W; Konzak, CF; von Wettstein, D; Rustgi, S |
| Journal | PLoS ONE |
| Volume | 8 |
| Issue | 11 |
| Page Numbers | e80155 |
| Abstract | Microspores can be induced to develop homozygous doubled haploid plants in a single generation. In the present experiments androgenic microspores of wheat have been genetically transformed and developed into mature homozygous transgenic plants. Two different transformation techniques were investigated, one employing electroporation and the other co-cultivation with Agrobacterium tumefaciens. Different tissue culture and transfection conditions were tested on nine different wheat cultivars using four different constructs. A total of 19 fertile transformants in five genotypes from four market classes of common wheat were recovered by the two procedures. PCR followed by DNA sequencing of the products, Southern blot analyses and bio/histo-chemical and histological assays of the recombinant enzymes confirmed the presence of the transgenes in the T0 transformants and their stable inheritance in homozygous T1∶2 doubled haploid progenies. Several decisive factors determining the transformation and regeneration efficiency with the two procedures were determined: (i) pretreatment of immature spikes with CuSO4 solution (500 mg/L) at 4°C for 10 days; (ii) electroporation of plasmid DNA in enlarged microspores by a single pulse of ∼375 V; (iii) induction of microspores after transfection at 28°C in NPB-99 medium and regeneration at 26°C in MMS5 medium; (iv) co-cultivation with Agrobacterium AGL-1 cells for transfer of plasmid T-DNA into microspores at day 0 for <24 hours; and (v) elimination of AGL-1 cells after co-cultivation with timentin (200-400 mg/L). |
| Doi | 10.1371/journal.pone.0080155 |
| Pmid | 24260351 |
| Wosid | WOS:000327308500118 |
| Is Certified Translation | No |
| Dupe Override | No |
| Is Public | Yes |
| Language Text | English |