We have developed a nove1 and reproducible system for recovery of fertile transgenic maize (Zea mays L.) plants. The transformation was performed using microprojectile bombardment of cultured shoot apices of maize with a plasmid carrying two linked genes, the Strepfomyces bygroscopicus phosphinothricin acetyltransferase gene (bar) and the potato proteinase inhibitor II gene, either alone or in combination with another plasmid containing the 5' region of the rice actin 1 gene fused to the Escbericbia coli p-glucuronidase gene (gus). Bombarded shoot apices were subsequently multiplied and selected under 3 to 5 mg/L glufosinate ammonium. Co-transformation frequency was 100% (146/146) for linked genes and 80% (41/51) for unlinked genes. Co-expression frequency of the bar and gus genes was 5 7 % (29/51). The co-integration, co-inheritance, and co-expression of bar, the potato proteinase inhibitor II gene, and gus in transgenic R,, R,, and R, plants were confirmed. Localized expression of the actin 1-CUS protein in the R, and R, plants was extensively analyzed by histochemical and fluorometric assays.The shoot tip, or shoot apex, consists of the shoot apical meristem, a region in which lateral organ primordia form, a subapical region of cell enlargement, and severa1 leaf primordia (Steeves and Sussex, 1989). The meristem region contains apical initial cells and subepidermal cells from which the gametes are derived (Medford, 1992). Theoretically, there are two possibilities for recovering transgenic plants via transfer of DNA into the shoot apical meristem. One possibility is that transgenic progeny may be directly produced via transformation of the subepidermal germline cells followed by the development of a partially transgenic reproductive organ. In this case, the primary transformants will always be chimeric. An alternative possibility is to multiply transgenic apical meristem cells and/or germ-line cells, which can be reprogrammed in the developmental direction under in vitro conditions. Transgenic plants can be regenerated from these cells with or without selection. Our previous research on maize (Zea mays L.) morphogenesis demonstrated that the maize meristem is morphogenetically plastic and can be manipulated to produce multiple shoots, somatic embryos, tassels, or ears in a relatively genotype-independent manner by simple variation of in vitro culture conditions (Zhong et al., 1992a(Zhong et al., , 1992b. Based on this concept, we transformed maize meristems via microprojectile bombardment with a series of chimeric genes, including bar, pin2, and gus.In this paper, we report the efficient recovery of fertile transgenic maize plants via a shoot-multiplication system after microprojectile bombardment of shoot tips. Maize shoot apices were transformed with a plasmid incorporating bar driven by the CaMV 35s promoter and pin2 with the wound-inducible pin2 promoter (Fig. l), either alone or in combination with another plasmid containing gus driven by the 5' region of Actl (Fig. 1). The co-integration and co-inherita...

No abstract

No abstract

The 3' termini of RNA1 and RNA2 of blueberry leaf mottle virus (BBLMV) were cloned and the cDNA sequence of a portion of the putative polymerase gene, the complete coat protein (CP) gene, and the 3' non-coding regions was determined. The N terminus of the coat protein gene was precisely located by comparison with the amino acid sequence determined by the Edman degradation sequencing of the purified coat protein. The coat protein gene encoded a polypeptide of 521 amino acids with a predicted M(r) of 57,542. Homology to BBLMV coat protein was highest with tomato ringspot virus (TomRSV) and cherry leaf roll virus (CLRV); two other nepoviruses also belonging to a sub-group defined by the presence of large RNA2 components. The 3' terminal 1390 nt of RNA1 and RNA2 were nearly identical and apparently non-coding. No statistically significant sequence homology was found between the 3' non-coding region of this length is unusual, but has been reported for two other related viruses, TomRSV and CLRV. The biological function of the long 3' non-coding region and how the high level of sequence homology is maintained between RNA1 and RNA2, is unknown. Possible mechanisms for conservation of the 3' terminus are discussed.

No abstract