An RFLP linkage map for loblolly pine based on a three-generation outbred pedigree

A genetic linkage map for loblolly pine (Pinus taeda L.) was constructed using segregation data from a three-generation outbred pedigree consisting of four grandparents, two parents, and 95 F₂ progeny. The map was based predominantly on restriction fragment length polymorphism (RFLP) loci detected by cDNA probes. Sixty-five cDNA and three genomic DNA probes revealed 90 RFLP loci. Six polymorphic isozyme loci were also scored. One-fourth (24%) of the cDNA probes detected more than 1 segregating locus, an indication that multigene families are common in pines. As many as six alleles were observed at a single segregating locus among grandparents and it was not unusual for the progeny to segregate for three or four alleles per locus. Multipoint linkage analysis placed 73 RFLP and 2 isozyme loci into 20 linkage groups; the remaining 17 RFLP and 4 isozyme loci were unlinked. The mapped RFLP probes provide a new set of codominant markers for genetic analyses in loblolly pine.     

Comparison of allozyme,RFLP, and RAPD markers for revealing genetic variation within and between trembling aspen and bigtooth aspen

We examined genetic variation in allozyme loci, nuclear DNA restriction fragment length polymorphisms (RFLPs), and random amplified polymorphic DNAs (RAPDs) in 130 trembling aspen (Populus tremuloides) and 105 bigtooth aspen (P. grandidentata) trees. In trembling aspen 10 out of 13 allozyme loci assayed (77%) were polymorphic (P), with 2.8 alleles per locus (A) and an expected heterozygosity (H_e) of 0.25. In contrast, bigtooth aspen had a much lower allozyme genetic variability (P=29%; A=1.4; H_e=0.08). The two species could be distinguished by mutually exclusive alleles at Idh-1, and bigtooth aspen has what appears to be a duplicate 6PG locus not present in trembling aspen. We used 138 random aspen genomic probes to reveal RFLPs in HindIII digests of aspen DNA. The majority of the probes were from sequences of low copy number. RFLP results were consistent with those of the allozyme analyses, with trembling aspen displaying higher genetic variation than bigtooth aspen (P=71%, A=2.7, and H_e=0.25 for trembling aspen; P=65%, A=1.8, and H_e=0.13 for bigtooth aspen). The two species could be distinguished by RFLPs revealed by 21 probes (15% of total probes assayed). RAPD patterns in both species were studied using four arbitrary decamer primers that revealed a total of 61 different amplified DNA fragments in trembling aspen and 56 in bigtooth aspen. Assuming a Hardy-Weinberg equilibrium, estimates of P=100%, A=2, and H_e=0.30 in trembling aspen and P=88%, A=1.9, and H_e=0.31 in bigtooth aspen were obtained from the RAPD data. Five amplified DNA fragments were species diagnostic. All individuals within both species, except for 2 that likely belong to the same clone, could be distinguished by comparing their RAPD patterns. These results indicate that (1) RFLPs and allozymes reveal comparable patterns of genetic variation in the two species, (2) trembling aspen is more genetically variable than bigtooth aspen at both the allozyme and DNA levels, (3) one can generate more polymorphic and species-specific loci with DNA markers than with allozymes in aspen, and (4) RAPDs provide a very powerful tool for fingerprinting aspen individuals.     

Mapping of a Magnaporthe grisea locus affecting rice (Oryza sativa) cultivar specificity

Magnaporthe grisea causes rice blast, the most important fungal disease of rice. The segregation of genes controlling virulence of M. grisea on rice was studied to establish the genetic basis of cultivar specificity in this host-parasite interaction. Full-sib progeny and parent isolates Guy11 and 2539 of M. grisea were inoculated onto rice (Oryza sativa) cultivar CO39 and five near-isogenic lines (NILs) of CO39. Each NIL contained a different single gene affecting resistance to specific isolates of M. grisea. No differential interactions between NILs and progeny or parents were observed; parents and progeny pathogenic on CO39 were pathogenic on all five NILs. Segregation ratios of 101 full-sib progeny, 117 progeny from full-sib parents, and 109 backcross progeny, indicated a common single gene affecting pathogenicity on CO39 and the five NILs. A subset of the above 327 isolates (43 fullsib progeny, 37 progeny from full-sib parents, and 32 backcross progeny) were inoculated onto rice cultivar 51583; all were pathogenic, indicating that cultivar specificity to CO39 was segregating in this population of isolates. The locus controlling cultivar specificity, named avrCO39, was mapped to chromosome 1 using a subset of the progeny previously used to construct an RFLP map of M. grisea. The closest reported RFLP markers were 11.8 (estimated 260 kb) and 17.2 cM (estimated 380 kb) away and provide starting points on either side of the locus for a chromosome walk to clone the locus.     

Dominance and recessiveness at loci for virulence against potato and tomato in Phytophthora infestans

Summary In this study we investigated the genetic control of virulence in the diploid fungal pathogen, Phytophthora infestans, against host resistance genes R1, R2, R3, and R4 (potato) and Ph1 (tomato). For four of these virulence traits, the presence or absence of segregation indicated conclusively which phenotype was dominant. We observed a 31 (virulentavirulent) segregation on R2 in the progeny of parents which were both virulent, suggesting that virulence is dominant and both parents are heterozygous. In a cross in which one parent was virulent and the other avirulent on potato gene R3, all progeny tested were avirulent, so avirulence against R3 is dominant. The same virulent parent crossed with a different avirulent parent produced virulent and avirulent progeny in a 13 ratio, indicating that a second locus may be involved. The progeny of two parents virulent on R4 segregated for virulence and avirulence, so virulence against R4 is dominant. For Ph1, a 13 segregation in the progeny of two avirulent parents showed that the avirulent phenotype is dominant, and a 31 ration in a second cross suggested the involvement of a second locus. The segregations for virulence against R1 did not indicate which phenotype was dominant, but did suggest singlelocus control.     

Inheritance of chloroplast and mitochondrial DNA in Picea and composition of hybrids from introgression zones

Summary The cloning of white spruce (Picea glauca) mitochondrial DNA homologous to the cytochrome oxidase II and ATPase genes of maize is described. These probes were used to define restriction fragment length polymorphisms which distinguish the white, Engelmann (P. engelmannii) and Sitka spruce (P. sitchensis) populations that occur in British Columbia. Analysis of progeny from crosses between the species revealed that mitochondrial DNA was maternally inherited in all cases (32 progeny from five independent crosses). The inheritance of chloroplast DNA was determined using a probe described previously; in this case, all progeny exhibited paternal inheritance (27 progeny from four crosses). Mitochondrial and chloroplast probes were used to test trees from zones of introgression between coastal (Sitka) and interior spruces (white and Engelmann). In most cases mitochondria and chloroplasts within individuals were contributed by different species. The data shows that there is a significant Sitka spruce component in trees east of the coastal watershed in British Columbia.     

Growth form distribution and genetic relationships in tree clusters of Pinus flexilis,a bird-dispersed pine

Seed dispersal by the Clark's nutcracker (Nucifraga columbiana Wilson) may markedly influence the growth form and genetic population structure of limber pine (Pinus flexilis James). The nutcracker buries clusters of seeds in subterranean caches; germination of clustered seeds often results in a growth form characterized by two or more genetically distinct trees with fused or contiguous trunks (tree clusters). The occurrence of a morphologically similar form, the multi-trunk tree (a single genet branched near the base), as well as the typical single-trunked tree, complicates the study of limber pine populations. We examined growth form distribution and genetic relationships in tree clusters in limber pine populations at four elevations (from 2585 m to 3460 m) in the Colorado Front Range. At three study areas, relative occurrence of limber pine growth forms, as well as that of associated pines, was examined by a point-centered quarter survey. From the four study areas, we collected foliage from each trunk from a total of 74 clumps (combined tree clusters and multi-trunk trees) in order to differentiate the two growth forms using starch gel protein electrophoresis. Tree clumps were significantly more common in limber pine than in ponderosa or lodgepole pine (P<0.010). Although single-trunk limber pine was the most common growth form, except at the highest elevation, both multi-trunk trees and tree clusters were present in each stand. Tree clusters were estimated to comprise about 20% of the tree sites in each limber pine stand; the estimated proportion of multi-trunk trees varied by site from 5% to 77%. Trees in clusters were related, on average, as half to full siblings (mean r=0.43), but were unrelated to trees in other clusters (mean r=0.01). Electrophoretic analysis suggests possible genetic differentiation in limber pine that may be the result of different selection pressures on the growth forms.     

High variability and disomic segregation of microsatellites in the octoploid Fragaria virginiana Mill. (Rosaceae)

The objectives of the present study were to develop microsatellite markers for the wild strawberry, Fragaria virginiana, to evaluate segregation patterns of microsatellite alleles in this octoploid species, and assess genetic variability at microsatellite loci in a wild population. A genomic library was screened for microsatellite repeats and several PCR primers were designed and tested. We also tested the use of heterologous primers and found that F. virginiana primers amplified products in cultivated strawberry, Fragaria × ananassa Duch. and Fragaria chiloensis. Similarly, microsatellite loci developed from cultivated strawberry also successfully amplified F. virginiana loci. We investigated four microsatellite loci in detail, three developed from F. virginiana and one from cultivated strawberry. A survey of 100 individuals from a population of F. virginiana in Pennsylvania demonstrated high heterozygosities (H_e or gene diversity ranged from 0.80 to 0.88 per locus) and allelic diversity (12–17 alleles per locus), but individual plants had no more than two alleles per locus. Segregation patterns in parents and progeny of two controlled crosses at these four loci were consistent with disomic Mendelian inheritance. Together these findings suggest that the genome of F. virginiana is "highly diploidized" and at least a subset of microsatellite loci can be treated as codominant, diploid markers. Significant heterozygote deficiencies were found at three of the four loci for hermaphroditic individuals but for only one locus among females in this gynodioecious species.Communicated by J. Dvorak     

Hypomethylated sequences: Characterization of the duplicate soybean genome

Soybean is believed to be a diploidized tetraploid generated from an allotetraploid ancestor. In this study, we used hypomethylated genomic DNA as a source of probes to investigate the genomic structure and methylation patterns of duplicated sequences. Forty-five genomic clones from Phaseolus vulgaris and 664 genomic clones from Glycine max were used to examine the duplicated regions in the soybean genome. Southern analysis of genomic DNA using probes from both sources revealed that greater than 15% of the hypomethylated genomic regions were only present once in the soybean genome. The remaining ca. 85% of the hypomethylated regions comprise duplicated or middle repetitive DNA sequences. If only the ratio of single to duplicate probe patterns is considered, it appears that 25% of the single-copy sequences have been lost. By using a subset of probes that only detected duplicated sequences, we examined the methylation status of the homeologous genomes with the restriction enzymes MspI and HpaII. We found that in all cases both copies of these regions were hypomethylated, although there were examples of low-level methylation. It appears that duplicate sequences are being eliminated in the diploidization process. Our data reveal no evidence that duplicated sequences are being silenced by inactivation correlated with methylation patterns.     

Inheritance of restriction fragment length polymorphisms and random amplified polymorphic DNAs in coastal Douglas-fir

A total of 225 new genetic loci [151 restriction fragment length polymorphisms (RFLP) and 74 random amplified polymorphic DNAs (RAPD)] in coastal Douglas-fir [Pseudotsuga menziesii (Mirb.) Franco var. menziesii] have been identified using a three-generation outbred pedigree. The Mendelian inheritance of 16 RFLP loci and 29 RAPD loci was demonstrated based on single-locus segregation in a sample of F₂ progeny. One RFLP locus, PtIFG2025, showed segregation distortion. Probe pPtIFG2025 is a loblolly pine cDNA probe encoding for rbcS. The 16 RFLP loci and 23 allozyme loci were also assayed in a sample of 16 Douglas-fir seed-orchard clones. Allelism was determined at 11 of the 16 RFLP loci. RFLPs were able to detect slightly more variation (4.0 alleles per locus) than allozymes (3.1 alleles per locus). The inheritance of an additional 80 RAPD loci was determined based on haploid segregation analysis of megagametophytes from parent tree 013-1. Once 200–300 markers are identified and placed on a genetic map, quantitative trait loci affecting bud phenology will be mapped.     

Morphological and molecular characterization of fertile tetraploid somatic hybrids produced by protoplast electrofusion and PEG-induced fusion between Lycopersicon esculentum Mill. and Lycopersicon peruvianum Mill.

Summary Mesophyl protoplasts of two genotypes of cultivated tomato (Lycopersicon esculentum Mill.) and one of its wild relative species (Lycopersicon peruvianum Mill.) were fused by using electrofusion and polyethyleneglycol-induced fusion. Forty-three fertile tetraploid somatic hybrid plants, each deriving from separate calli, were recovered from both fusion procedures. Electrofusion appeared more efficient than chemical fusion for the production of somatic hybrids. These plants appeared morphologically similar, whatever the fusion procedure and tomato genotype. They had intermediate leaf, inflorescence, and flower morphology. After self-pollination, the hybrids set fruit of intermediate size and color. The hybrid nature of these plants was confirmed by isoelectric focusing of the Rubisco small subunits used as nuclear markers. L. esculentum and L. peruvianum were distinguished by means of two chloroplast markers: CF1-ATPase subunit as analyzed by isoelectro-focusing and ct DNA restriction patterns. All hybrids displayed both ct markers of only one parent with no biased transmission. Mitochondrial (mt) DNAs were prepared from flower buds by using miniaturized CsCl gradients. Preliminary analysis indicated that mt genomes from the hybrids all differed from those of both parents. mt DNA Sall restriction enzyme analysis revealed that all but two hybrids contained one novel fragment of 13.5 kb. Gene mapping experiments showed that the mt apocytochrome b and ATPase subunit 9 homologies in the somatic hybrid mt DNA resembled L. esculentum and L. peruvianum, respectively; the mt nad5 probe distinguished at least four distinct patterns in the hybrids. These results indicated that mt DNA rearrangements involving intergenomic recombinations occurred through protoplast fusion. A greater mt DNA polymorphism was induced with chemical fusion than with electrofusion.     

Leaf-rust resistance in rye (<Emphasis Type="Italic">Secale cereale</Emphasis> L.). 2. Genetic analysis and mapping of resistance genes <Emphasis Type="Italic">Pr3</Emphasis>, <Emphasis Type="Italic">Pr4</Emphasis>, and <Emphasis Type="Italic">Pr5</Emphasis>

The conventional way to drive modifications in old forest tree seed orchards is to establish progeny trials involving each parent tree and then evaluate its contribution to the performance of the progeny by estimating its general and specific combining ability (GCA and SCA). In this work, we successfully applied an alternative parent selection tactic based on paternity testing of superior offspring derived from a hybrid seed orchard established with a single Eucalyptus grandis seed parents and six E. urophylla pollen parents. A battery of 14 microsatellite markers was used to carry out parentage tests of 256 progeny individuals including two independent samples of selected trees and one control unselected sample, all derived from 6-year-old forest stands in eastern Brazil. Paternity determination was carried out for all progeny individuals by a sequential paternity exclusion procedure. Exclusion was declared only when the obligatory paternal allele in the progeny tree was not present in the alleged parent tree for at least four independent markers to avoid false exclusions due to mutation or null alleles. After maternity checks to identify seed mixtures and selfed individuals, the paternity tests revealed that approximately 29% of the offspring was sired by pollen parents outside the orchard. No selfed progeny were found in the selected samples. Three pollen parents were found to have sired essentially all of the offspring in the samples of selected and non-selected progeny individuals. One of these three parents sired significantly more selected progeny than unselected ones (P0.0002 in a Fisher exact test). Based on these results, low-reproductive-successful parents were culled from the orchard, and management procedures were adopted to minimize external pollen contamination. A significant difference (P<0.01) in mean annual increment was observed between forest stands produced with seed from the orchard before and after selection of parents and revitalization of the orchard. An average realized gain of 24.3% in volume growth was obtained from the selection of parents as measured in forest stands at age 2–4 years. The marker-assisted tree-breeding tactic presented herein efficiently identified top parents in a seed orchard and resulted in an improved seed variety. It should be applicable for rapidly improving the output quality of seed orchards, especially when an emergency demand for improved seed is faced by the breeder.Communicated by D.B. Neale     

Use of RFLP markers for the identification of alleles of the Pm3 locus conferring powdery mildew resistance in wheat (Triticum aestivum L.)

The objective of this study was to identify molecular markers linked to genes for resistance to powdery mildew (Pm) in wheat using a series of Chancellor near-isogenic-lines (NILs), each having one powdery mildew resistance gene. A total of 210 probes were screened for their ability to detect polymorphism between the NILs and the recurrent parent. One of these restriction fragment length polymorphism (RFLP) markers (Xwhs179) revealed polymorphism not only between the NILs for the Pm3 locus, but also among NILs possessing different alleles of the Pm3 locus. The location of the marker Xwhs179 was confirmed to be on homoeologous chromosome group 1 with the help of nullitetrasomic wheat lines. The linkage relationship between this probe and the Pm3 locus was estimated with double haploid lines derived from a cross between wheat cvs Club and Chul (Pm3b). The genetic distance was determined to be 3.3±1.9 cM.     

A genetic linkage map for Pinus radiata based on RFLP, RAPD, and microsatellite markers

A genetic linkage map for radiata pine (Pinus radiata D. Don) has been constructed using segregation data from a three-generation outbred pedigree. A total of 208 loci were analyzed including 165 restriction fragment length polymorphism (RFLP), 41 random amplified polymorphic DNA (RAPD) and 2 microsatellite markers. The markers were assembled into 22 linkage groups of 2 or more loci and covered a total distance of 1382 cM. Thirteen loci were unlinked to any other marker. Of the RFLP loci that were mapped, 93 were detected by loblolly pine (P. taeda L.) cDNA probes that had been previously mapped or evaluated in that species. The remaining 72 RFLP loci were detected by radiata pine probes from a PstI genomic DNA library. Two hundred and eighty RAPD primers were evaluated, and 41 loci which were segregating in a 11 ratio were mapped. Two microsatellite markers were also placed on the map. This map and the markers derived from it will have wide applicability to genetic studies in P. radiata and other pine species.     

Production and genetic characterization of near-isogenic lines in the bread-wheat cultivar Alpe

Two biotypes of the bread-wheat cultivar Alpe were shown to possess contrasting alleles at each of the glutenin (Glu-B1, Glu-D1, Glu-B3 and Glu-D3) and gliadin (Gli-B1 and Gli-D1) loci on chromosomes 1B and 1D. Fourteen near-isogenic lines (NILs) were produced by crossing these biotypes and used to determine the genetic control of both low-molecular-weight (LMW) glutenin subunits and gliadins by means of one-dimensional or two-dimensional electrophoresis. Genes coding for the B, C and D groups of EMW subunits were found to be inherited in clusters tightly linked with those controlling gliadins. Southern-blot analysis of total genomic DNAs hybridized to a -gliadin-specific cDNA clone revealed that seven NILs lack both the Gli-D1 and Glu-D3 loci on chromosome 1D. Segregation data indicated that these null alleles are normally inherited. Comparison of the null NILs with those possessing allele b at the Glu-D3 locus showed one B subunit, seven C subunits and two D subunits, as fractionated by two-dimensional A-PAGExSDS-PAGE, to be encoded by this allele. Alleles b and k at Glu-B3 were found to code for two C subunits plus eight and six B subunits respectively, whereas alleles b and k at Gli-B1 each controlled the synthesis of two -gliadins, one and two -gliadins. The novel Gli-B5 locus coding for two -gliadins was shown to recombine with the Gli-B1 locus on chromosome 1B. The two-dimensional map of glutenin subunits showed -gliadins encoded at the Gli-A2 locus on chromosome 6A. The use of Alpe NILs in the study of the individual and combined effects of glutenin subunits on dough properties is discussed.Research supported by a grant from the Commission of the European Communities, ECLAIR programme, Contract AGRE 0052     

The R1 gene conferring race-specific resistance to Phytophthora infestans in potato is located on potato chromosome V.

Summary Late blight in potato is caused by the fungusPhytophthora infestans and can inflict severe damage on the potato crop. Resistance toP. infestans is either based on major dominantR genes conferring vertical, race-specific resistance or on minor genes inducing horizontal, unspecific resistance. A dihaploid potato line was identified which carried theR1 gene, conferring vertical resistance to allP. infestans races, with the exception of those homozygous for the recessive virulence allele of the locusV1. The F₁ progeny of a cross between this resistant parent P(R1) and P(r), a line susceptible to all races, was analysed for segregation ofR1 and of restriction fragment length polymorphism (RFLP) markers distributed on the potato RFLP map comprising more than 300 loci. TheR1 locus was mapped to chromosome V in the interval between RFLP markers GP21 and GP179. The map position ofR1 was found to be very similar to the one ofRx2, a dominant locus inducing extreme resistance to potato virus X.     

Quantitative genetics of growth and development in Populus. I. A three-generation comparison of tree architecture during the first 2 years of growth

One approach to gain an insight into the genetics of tree architecture is to make use of morphologically divergent parents and study their segregating progeny in the F₂ and backcross (B₁) generations. This approach was chosen in the present study in which material of a three-generation pedigree growing side by side in a replicated plantation, was analyzed. The pedigree included Populus trichocarpa (T) and P. deltoides (D) parents, their F₁ and F₂ hybrids and their B₁ hybrids to the D parent. The trees were grown in the environment of the T parent and measured for the first 2 years of growth. Nine quantitative traits were studied at the stem, branch and leaf levels of tree architecture, in which the original parents differed. Strong F₁ hybrid vigor relative to the better parent (T) was expressed in growth and its components. Most quantitative traits in the F₂ and B₁ hybrids were intermediate between the T and D parents but displayed a wide range of variation due to segregation. The results from the analysis of variance indicated that all morphometric traits were significantly different among F₂ and B₁ clones, but the B₁ hybrids were more sensitive to replicates than the F₂. Broad-sense heritabilities (H ²) based on clonal means ranged from moderately high to high (0.50–0.90) for the traits studied, with H ² values varying over age. The H ² estimates reflected greater environmental noise in the B₁ than in the F₂, presumably due to the greater proportion of maladaptive D alleles in those hybrids. In both families, sylleptic branch number and length, and leaf size on the terminal, showed strong genetic correlations with stem growth. The large divergence between the two original parents in the traits studied, combined with the high chromosome number in Populus (2n=38), makes this pedigree well suited for the estimation of the number of quantitative trait loci (QTLs) underlying quantitative variation by Wright's biometric method (1968). Variation in several traits was found to be under the control of surprisingly few major QTLs: 3–4 in 2nd-year height and diameter growth, a single QTL in stem diameter/height ratio.     

A somaclonal variant of wheat with additional β-amylase isozymes

Summary The progeny of 149 plants regenerated from tissue culture of immature wheat (Triticum aestivum) embryos were screened for variation in their grain -amylase isozyme pattern. One regenerant was found which was heterozygous for a variant pattern characterized by the presence of at least five new isozyme bands, as well as an increased intensity in existing bands in two more positions. The F₂ of a homozygous variant crossed back to the parent segregated in an approximate 31 ratio but resolution of the gels was not sufficient to distinguish whether this represents a dominant or co-dominant single mutant gene. No chromosome abnormalities were evident in mitosis or meiosis of the homozygous variant or in the F₁ of the variant crossed back to the parent. No recombination has been seen between the variant bands and production of multiple bands from a single locus is consistent with the nature of the known -amylase loci. However, the variant bands were not evident in a survey of 111 diverse genotypes, nor were they present in developing grain of the parent cultivar. Therefore, this variant could represent a rare mutation leading to expression of a currently unexpressed locus.     

Identification of RFLP markers linked to the cereal cyst nematode resistance gene (Cre) in wheat

The cereal cyst nematode (CCN) (Heterodera avenae Woll.) is an economically damaging pest of wheat in many of the worlds cereal growing areas. The development of CCN-resistant cultivars may be accelerated by the use of molecular markers. The Cre gene of the wheat line AUS 10894 confers resistance to CCN. Using a pair of near-isogenic lines (NILs) that should differ only in a small chromosome segment containing the Cre locus, we screened 58 group-2 probes and found two (Tag605 and CDO588) that detect polymorphism between the NILs. Nulli-tetrasomic and ditelosomic lines confirmed that the restriction fragment length polymorphism (RFLP) markers identified were derived from the long arm of wheat chromosome 2. Crosses between AUS 10894 and Spear and the NIL AP and its recurrent parent Prins were used to produce F₂ populations that gave the expected 31 segregation ratio for the resistance gene. Linkage analysis identified two RFLP markers flanking the resistance gene. Xglk605 and Xcdo588 mapped 7.3 cM (LOD=6.0) and 8.4 cM (LOD=6.7), respectively, from the Cre locus.     

Directed excision of a transgene from the plant genome

Summary The effectiveness of loxP-Cre directed excision of a transgene was examined using phenotypic and molecular analyses. Two methods of combining the elements of this system, re-transformation and cross pollination, were found to produce different degrees of excision in the resulting plants. Two linked traits, -glucuronidase (GUS) and a gene encoding sulfonylurea-resistant acetolactate synthase (ALS^r), were integrated into the genome of tobacco and Arabidopsis. The ALS^r gene, bounded by loxP sites, was used as the selectable marker for transformation. The directed loss of the ALS^T gene through Cre-mediated excision was demonstrated by the loss of resistance to sulfonylurea herbicides and by Southern blot analysis. The -glucuronidase gene remained active. The excision efficiency varied in F₁ progeny of different lox and Cre parents and was correlated with the Cre parent. Many of the lox × Cre F₁ progeny were chimeric and some F₂ progeny retained resistance to sulfonylureas. Re-transformation of lox/ALS/lox/GUS tobacco plants with cre led to much higher efficiency of excision. Lines of tobacco transformants carrying the GUS gene but producing only sulfonylurea-sensitive progeny were obtained using both approaches for introducing cre. Similarly, Arabidopsis lines with GUS activity but no sulfonylurea resistance were generated using cross pollinations.