Identification of Brassica oleracea monosomic alien chromosome addition lines with molecular markers reveals extensive gene duplication

Summary Chromosomes of Brassica oleracea (2n=18) were dissected from the resynthesized amphidiploid B. napus Hakuran by repeated backcrosses to B. campestris (2n=20), creating a series of monosomic alien chromosome addition line plants (2n=21). Using morphological, isozyme and restriction fragment length polymorphism markers (RFLPs), 81 putative loci were identified. Of nine possible synteny groups, seven were represented in the 25 monosomic addition plants tested. Sequences homologous to 26% of the 61 DNA clones utilized (80% were cDNA clones) were found on more than one synteny group, indicating a high level of gene duplication. Anomalous synteny associations were detected in four 2n=21 plants. One of these plants showed two markers from one B. oleracea chromosome associated with a second complete B. oleracea synteny group, suggesting translocation or recombination between non-homologous chromosomes in Hakuran or the backcross derivatives. The other three 2n=21 plants each contained two or more B. oleracea synteny groups, suggesting chromosome substitution.     

Integrated map of AFLP, SSLP and RFLP markers using a recombinant inbred population of rice (Oryza sativa L.)

 A molecular map of rice consisting of 231 amplified fragment length polymorphisms (AFLPs), 212 restriction fragment length polymorphisms (RFLPs), 86 simple-sequence length polymorphisms (SSLPs), five isozyme loci, and two morphological mutant loci [phenol staining of grain (Ph), semi-dwarf habit (sd-1)] has been constructed using an F₁₁ recombinant inbred (RI) population. The mapping population consisted of 164 RI lines and was developed via single-seed descent from an intercross between the genetically divergent parents Milyang 23 (M) (tongil type) and Gihobyeo (G) ( japonica type). A subset of previously mapped RFLP and SSLP markers were used to construct the map framework. The AFLP markers were derived from ten EcoRI(+2) and MseI(+3) primer combinations. All marker types were well distributed throughout the 12 chromosomes. The integrated map covered 1814 cM, with an average interval size of 3.4 cM. The MG map is a cornerstone of the Korean Rice Genome Research Program (KRGRP) and is being continuously refined through the addition of partially sequenced cDNA markers derived from an immature-seed cDNA library developed in Korea, and microsatellite markers developed at Cornell. The population is also being used for quantitative trait locus (QTL) analysis and as the basis for marker-assisted variety development. Received: 24 June 1997 / Accepted: 25 November 1997     

Generation and characterisation of monosomic chromosome addition lines of Brassica campestris - B. oxyrrhina

 Monosomic chromosome addition lines of Brassica oxyrrhina in the background of alloplasmic B. campestris carrying B. oxyrrhina cytoplasm were generated and characterised through morphology, cytology and molecular (RAPD) analysis. Four successive backcrosses of the synthetic alloploid B. oxycamp with B. campestris yielded 24 monosomic addition plants that were grouped into seven different synteny groups based on morphological similarity and RAPD patterns. Each synteny group exhibited morphological features diagnostic for the presence of individual B. oxyrrhina chromosomes including some novel phenotypes. Meiotic studies of the addition lines revealed the homoeology of four B. oxyrrhina chromosomes (synteny groups 1, 3, 5 and 6 ) with B. campestris chromosomes as indicated by trivalent associations, with the highest homoeology (44.23%) in synteny group 1 and the lowest (6.1%) in synteny group 3. Seed fertility of the addition lines ranged from 94.85% (synteny group 1) to 56.98% (synteny group 5). All of the addition lines were male-sterile except synteny group 6 which had 12–16% stainable pollen. Ovule transmission of the B. oxyrrhina chromosomes added to the progenies of addition lines ranged from 23.52% (synteny group 6) to 14% (synteny group 7). RAPD analysis confirmed the validity of synteny grouping based on morphological observations. Approximately 45% of the primers studied were informative, giving B. oxyrrhina-specific RAPD bands unique for each synteny group, except group 6. Received: 20 October 1997 / Accepted: 31 March 1998     

Mapping of DNA markers to arms and sub-arm regions of Nicotiana sylvestris chromosomes using aberrant alien addition lines

Seven monosomic addition plants, each containing the full complement of Nicotiana plumbaginifolia (2n = 20, genome constitution PP) and an aberrant chromosome of Nicotiana sylvestris (2n = 24, SS), were produced from backcrosses of hyperdiploid derivatives of the sesquidiploid hybrid PPS to N. plumbaginifolia. The N. sylvestris chromosomes in these plants were characterized by karyotype analysis, Southern hybridization with DNA markers previously localized on N. sylvestris chromosomes and a 269-bp fragment from the 3' end of 25S rDNA, and fluorescence in situ hybridization using 25S rDNA, 5S rDNA and telomere repeats (TTTAGGG)_n as probes. The N. sylvestris chromosomes in these plants were identified to be telocentrics 6S, 7S and 8S, and deletions 7S, 10, 12S and 12L, respectively. The successful identification of aberrant chromosomes in these lines enabled us to assign DNA markers to arms and sub-arm regions of N. sylvestris chromosomes. All aberrant chromosomes in the addition lines could be transmitted through mitosis and meiosis. The potential applications of the addition lines in high-resolution physical mapping, the isolation of N. sylvestris chromosomes by flow cytometry, and an understanding of the chromosomal distribution of 45S rDNA in N. sylvestris are discussed.     

Comparison of the genetic diversity of common wild rice (Oryza rufipogon Griff.) and cultivated rice (O. sativa L.) using RFLP markers

Forty fourth single-copy RFLP markers were used to evaluate the genetic diversity of 122 accessions of common wild rice (CWR, Oryza rufipogon Griff.) and 75 entries of cultivated rice (Oryza sativa L. ) from more than ten Asian countries. A comparison of the parameters showing genetic diversity, including the percentage of polymorphic loci (P), the average number of alleles per locus (A), the number of genotypes (Ng), the average heterozygosity (Ho) and the average genetic multiplicity (Hs) of CWR and indica and japonica subspecies of cultivated rice from different countries and regions, indicated that CWR from China possesses the highest genetic diversity, followed by CWR from South Asia and Southeast Asia. The genetic diversity of CWR from India is the second highest. Although the average gene diversity (Hs)of the South Asian CWR is higher than that of the Southeast Asian CWR, its percentage of polymorphic loci (P), number of alleles (Na) and number of genotypes (Ng) are all smaller. It was also found that the genetic diversity of cultivated rice is obviously lower than that of CWR. At the 44 loci investigated, the number of polymorphic loci of cultivated rice is only 3/4 that of CWR, while the number of alleles, 60%, and the number of genotypes is about 1/2 that of CWR. Of the two subspecies studied, the genetic diversity of indica is higher than that of japonica. The average heterozygosity of the Chinese CWR is the highest among all the entries studied. The average heterozygosity of CWR is about two-times that of cultivated rice. It is suggested that during the course of evolution from wild rice to cultivated rice, many alleles were lost through natural and human selection, leading to the lower heterozygosity and genetic diversity of the cultivated rice. Received: 19 May 1999 / Accepted: 26 April 2000     

AFLP linkage group assignment to the chromosomes of Allium cepa L. via monosomic addition lines

Two complete sets of Allium fistulosum L.– A. cepa monosomic addition lines (2n=2x+1=17) together with an AFLP linkage map based on a cross between A. cepa and A. roylei Stearn were used to re-evaluate the eight A. cepa linkage groups identified in the mapping study. The linkage groups could be assigned to individual, physical chromosomes. The low level of molecular homology between A. cepa and A. fistulosum enabled the identification of 186 amplified fragment length polymorphisms (AFLP™ markers) present in A. cepa and not in A. fistulosum with ten different primer combinations. With the monosomic addition lines the distribution of the markers over the eight chromosomes of A. cepa could be determined. Of these 186 AFLP markers 51 were absent in A. roylei and consequently used as markers in the mapping study (A. cepa ×A. roylei cross). Therefore, these 51 AFLP markers could be used to assign the eight A. cepa linkage groups identified in the mapping study to physical chromosomes. Seven isozyme and three CAPS markers were also included. Two of the linkage groups had to be split because they included two sets of markers corresponding to different chromosomes. A total of 20 (approx. 10%) of the A. cepa-specific AFLP markers were amplified in more than one type of the monosomic addition lines, suggesting unlinked duplications. The co-dominant isozyme and CAPS markers were used to identify the correspondence of linkage groupsoriginating from A. cepa or from A. roylei. Received: 16 April 1999 / Accepted: 13 August 1999     

An interspecific (Capsicum annuum ×C. chinese) F2 linkage map in pepper using RFLP and AFLP markers

We have constructed a molecular linkage map of pepper (Capsicum spp.) in an interspecific F₂ population of 107 plants with 150 RFLP and 430 AFLP markers. The resulting linkage map consists of 11 large (206–60.3 cM) and 5 small (32.6–10.3 cM) linkage groups covering 1,320 cM with an average map distance between framework markers of 7.5 cM. Most (80%) of the RFLP markers were pepper-derived clones, and these markers were evenly distributed across the genome. By using 30 primer combinations, we were able to generate 444 AFLP markers in the F₂ population. The majority of the AFLP markers clustered in each linkage group, although PstI/MseI markers were more evenly distributed than EcoRI/MseI markers within the linkage groups. Genes for the biosynthesis of carotenoids and capsaicinoids were mapped on our linkage map. This map will provide the basis of studying secondary metabolites in pepper. Received: 20 October 1999 / Accepted: 3 July 2000     

Establishment of a complete series of a monosomic tomato chromosome addition lines in the cultivated potato using RFLP and GISH analyses

With the aim of establishing a complete monosomic alien tomato chromosome addition series in a potato background, the backcross progenies derived from repeated crossing of potato (+) tomato fusion hybrids to potato were screened through RFLP and GISH analyses. Because of the availability from our previous work of seven of the possible 12 tomato monosomic additions, selected from BC₂ populations, attention was paid to those alien additions that were missing. Thus, since the alien additions were already available for tomato chromosomes 1, 2, 4, 6, 8, 10 and 12, efforts were made to select for chromosomes 3, 5, 7, 9 and 11 by screening specific BC₃ populations. In all, 105 plants from four BC₃ populations were screened through a combination of RFLP and GISH analyses in order to complete the series. Among the newly selected alien addition lines, five were monosomic additions for all the remaining chromosomes and one was a disomic addition for chromosome 11. When using conventional cytogenetics the selection of monosomic alien additions is highly laborious. All the tomato chromosomes showed a variable rate of transmission. Chromosome 6 was transmitted at 29.6% and 81.5% frequency in populations 2705 and 2701 respectively. The present study showed that molecular markers and molecular cytogenetics applied in this study were most efficient and rapid because a pre-selection for the desired genotypes was possible by screening a population with chromosome-specific markers for the presence of one tomato chromosome at a time. Received: 9 January 2001 / Accepted: 26 January 2001     

Differentiation and classification of parental lines and favorable genic interactions affecting F1 fertility in distant crosses of rice (Oryza sativa L.)

 This study was intended to investigate the extent of genetic differentiation in parental lines of rice hybrids and to analyze the genetic basis underlying the fertility phenomenon in distant crosses. Two subsets of rice material (111 entries in total) were used, including 81 doubled-haploid (DH) lines and 30 Indica and Japonica rice varieties or lines (as a control). The DH lines was derived from a heterotic Indica/Japonica cross (Gui630/02428) by anther culture. The materials in the control represent a broad spectrum of the Asian cultivated rice gene pool including landraces, primitive cultivars, historically important cultivars, modern elite cultivars, super rice and parents of superior hybrids. In accordance with the NC II design, 57 out of the DH lines were test-crossed to two important wide compatibility lines: photoperiod-sensitive genetic male sterile (PGMS) line N422s and thermo-sensitive genetic male sterile (TGMS) line Peiai64s. The F₁s and their parents, 182 entries in total, were examined for the performance of seven traits in a replicated field trial. All the rice materials was surveyed for polymorphisms using 92 RFLP markers selected from two published molecular marker linkage maps. Genotypes of the F₁ hybrids at the molecular-marker loci were deduced from the parental genotypes. The analysis showed that there were two types of genetic differentiation in the two subsets of rice material; that is, qualitative differentiation in the control and quantitative differentiation in the DH lines. In addition, favorable genic interactions (both intra- or inter-locus) contributed to better increase the fertility in hybrids of distant crosses through incorporation of a wide-compatibility line as the female parent. Favorable genic interactions can be applied in hybrid rice breeding programs by selecting parents with an appropriate extent of genetic differentiation. Received: 5 June 1997 / Accepted: 10 September 1997     

Assignment of RFLP linkage groups to chromosomes using monosomic F1 analysis in hexaploid oat

The availability of molecular genetic maps in oat (Avena spp.) and improved identification of chromosomes by C-banding are two recent developments that have made locating linkage groups to chromosomes possible in cultivated hexaploid oat, 2n=6x=42. Monosomic series derived from Avena byzantina C. Koch cv Kanota and from Avena sativa L. cv Sun II were used as maternal plants in crosses with the parents, Kanota-1 and Ogle-C, of the oat RFLP mapping population. Monosomic F₁ plants were identified by root-tip cell chromosome counts. For marker analysis, DNAs of eight F₂ plants from a monosomic F₁ were combined to provide a larger source of DNA that mimicked that of the monosomic F₁ plant. Absence of maternal alleles in monosomic F₁s served to associate linkage groups with individual chromosomes. Twenty two linkage groups were associated with 16 chromosomes. In seven instances, linkage groups that were independent of each other in recombination analyses were associated with the same chromosome. Five linkage groups were shown to be associated with translocation differences among oat lines. Additionally, the results better-characterized the oat monosomic series through the detection of duplicates and translocation differences among the various monosomic lines. The F₁ monosomic series represents a powerful cytogenetic tool with the potential to greatly improve understanding of the oat genome. Received: 24 April 2000 / Accepted: 10 May 2000     

Molecular mapping of genes conferring aluminum tolerance in rice (Oryza sativa L.)

Crop productivity on acid soil is restricted by multiple abiotic stress factors. Aluminum (Al) tolerance seems to be a key to productivity on soil with a pH below 5.0, but other factors such as Mn toxicity and the deficiency of P, Ca and Mg also play a role. The development of Al-tolerant genotypes of rice is an urgent necessity for improving crop productivity in developing countries. Inhibition of root growth is a primary and early symptom of Al toxicity. The present study was conducted to identify genetic factors controlling the aluminum tolerance of rice. Several parameters related to Al tolerance, most importantly the relative root growth under Al stress versus non-stress conditions, were scored in 188 F₃ selfed families from a cross between an Al-tolerant Vietnamese local variety, Chiembau, and an Al-susceptible improved variety, Omon269–65. The two varieties are both Oryza sativa ssp. indica, but showed a relatively high level of DNA polymorphism, permitting the assembly of an RFLP map consisting of 164 loci spanning 1,715.8 cM, and covering most of the rice genome. A total of nine different genomic regions on eight chromosomes have been implicated in the genetic control of root and shoot growth under aluminum stress. By far the greatest effects on aluminum tolerance were associated with the region near WG110 on chromosome 1. This region does not seem to correspond to most of the genes that have been mapped for aluminum tolerance in other species, nor do they correspond closely to one another. Most results, both from physiological studies and from molecular mapping studies, tend to suggest that aluminum tolerance is a complex multi-genic trait. The identification of DNA markers (such as WG110) that are diagnostic for aluminum tolerance in particular gene pools provides an important starting point for transferring and pyramiding genes that may contribute to the sustainable improvement of crop productivity in aluminum-rich soils. The isolation of genes responsible for aluminum tolerance is likely to be necessary to gain a comprehensive understanding of this complex trait. Received: 29 March 2000 / Accepted: 16 August 2000     

Development of a complete set of Triticum aestivum-Aegilops speltoides chromosome addition lines

Aegilops speltoides Tausch (2n = 2x = 14, SS) is considered as the closest living relative of the B and G genomes of polyploid wheats. A complete set of Triticum aestivum L. cv Chinese Spring-Ae. speltoides whole chromosomes and seven telosomic addition lines was established. A low pairing accession was selected for the isolation of the chromosome addition lines. Except for chromosomes 3S and 6S, which are presently only available as monosomic additions, all other lines were recovered as disomic or ditelosomic additions. The individual Ae. speltoides chromosomes isolated in the wheat background were assayed for their genetic effects on plant phenotype and cytologically characterized in terms of chromosome length, arm ratio, distribution of marker C-bands, and FISH sites using a Ae. speltoides-specific repetitive element, Gc1R-1, as a probe. The homoeology of the added Ae. speltoides chromosomes was established by using a standard set of RFLP probes. No chromosomal rearrangements relative to wheat were detected. Received: 28 June 1999 / Accepted: 16 November 1999     

Conversion of AFLP markers to sequence-specific PCR markers in barley and wheat

 Conversion of amplified fragment length polymorphisms (AFLPs) to sequence-specific PCR primers would be useful for many genetic-linkage applications. We examined 21 wheat nullitetrasomic stocks and five wheat-barley addition lines using 12 and 14 AFLP primer combinations, respectively. On average, 36.8% of the scored AFLP fragments in the wheat nullitetrasomic stocks and 22.3% in the wheat-barley addition lines could be mapped to specific chromosomes, providing approximately 461 chromosome-specific AFLP markers in the wheat nullitetrasomic stocks and 174 in the wheat-barley addition lines. Ten AFLP fragments specific to barley chromosomes and 16 AFLP fragments specific to wheat 3BS and 4BS chromosome arms were isolated from the polyacrylamide gels, re-amplified, cloned and sequenced. Primer sets were designed from these sequences. Amplification of wheat and barley genomic DNA using the barley derived primers revealed that three primer sets amplified DNA from the expected chromosome, five amplified fragments from all barley chromosomes but not from wheat, one amplified a similar-sized fragment from multiple barley chromosomes and from wheat, and one gave no amplification. Amplification of wheat genomic DNA using the wheat-derived primer sets revealed that three primer sets amplified a fragment from the expected chromosome, 11 primer sets amplified a similar-sized fragment from multiple chromosomes, and two gave no amplification. These experiments indicate that polymorphisms identified by AFLP are often not transferable to more sequence-specific PCR applications. Received: 30 June 1998 / Accepted: 26 October 1998     

Mapping of the QTL (quantitative trait locus) conferring partial resistance to leaf blast in rice cultivar Chubu 32

The rice cultivar Chubu 32 possesses a high level of partial resistance to leaf blast. The number and chromosomal location of genes conferring this resistance were detected by restriction fragment length polymorphism (RFLP) linkage mapping and quantitative trait locus (QTL) analysis. For the mapping, 149 F₃ lines derived from the cross between rice cultivar Norin 29, with a low level of partial resistance, and Chubu 32 were used, and their partial resistance to leaf blast was assessed in upland nurseries. A linkage map covering six chromosomes and consisting of 36 RFLP markers was constructed. In the map, only one significant QTL (LOD>2.0) for partial resistance was detected on chromosome 11. This QTL explained 45.6% of the phenotypic variation. The segregation ratio of the F3 lines was 3:1 for partial resistance to susceptibility. These results suggest that the partial resistance in Chubu 32 is controlled by a major gene. Received: 15 March 2001 / Accepted: 13 August 2001     

RFLP linkage map of the Ethiopian cereal tef [Eragrostis tef (Zucc) Trotter]

Tef [Eragrostis tef (Zucc)Trotter] is one of the most important cereal crops in Ethiopia. It is an allotetraploid species with a genome size of 720 Mbp. In this paper we report results of genetic linkage-map construction for E. tef using tef and heterologous cDNA probes for the first time. One hundred and sixteen F₈ recombinant inbred lines (RILs) from the cross E. tef cv Kaye Murri×Eragrostis pilosa (accession30–5) were used for mapping. Parental lines were digested with nine restriction enzymes and screened using 159 tef cDNA and 162 heterologous probes including the grass genome anchor probes. The polymorphism level between parental lines was 66.9%. One hundred and thirty nine polymorphic probes were hybridized against 116 RILs. Both the tef and the heterologous probes hybridized well against tef genomic DNA. The linkage map defined 1,489 cM of the tef genome comprising 149 marker loci distributed among 20 linkage groups. The average interval between markers was 9.99 cM. A fraction (14.8%) of the markers deviated significantly from the expected segregation. Such a genetic linkage map is useful for tagging economically useful genes in tef because a wide range of agronomically important traits is segregating within this population. This would enable the use of a marker assisted breeding strategy which, in turn, will enhance breeding efficiency. Alignment of the tef RFLP map with the rice RFLP map indicates that a number of syntenic chromosomal fragments exist between tef and rice in which the gene order was for the most part collinear. The comparative mapping information should enable tef scientists to take advantage of whatever genetic progress is made on the cereal model species rice. Received: 9 June 2000 / Accepted: 31 August 2000     

Analysis of mitochondrial DNA restriction fragment length polymorphisms for examining genetic variability among isolates of Phellinus linteus

 Restriction fragment length polymorphism (RFLP) analysis of mitochondrial DNAs (mtDNAs) from nine Japanese wild isolates of Phellinus linteus was carried out to examine their genetic variability. BamHI and EcoRI digests of mtDNAs from these isolates produced four and five distinct RFLP patterns, respectively. By combining the RFLP patterns obtained with the two endonucleases, mtDNAs from the nine isolates could be assigned to five different genotypes, but no mtDNA variation was detected among the isolates collected from a small area. Distance values calculated among all pairs of mtDNA genotypes, based on the presence or absence of comigrating restriction fragments, were clearly smaller than those among the mtDNA genotypes of Lentinula edodes and Pleurotus ostreatus samples collected worldwide, suggesting the necessity of collecting P. linteus wild isolates for genetic resources from geographically wider areas. Received: June 27, 2002 / Accepted: August 19, 2002 Correspondence to:T. Nakamura     

Telomere-homologous sequences occur near the centromeres of many tomato chromosomes

Several bacteriophage lambda clones containinginterstitialtelomererepeats (ITR) were isolated from a library of tomato genomic DNA by plaque hybridization with the clonedArabidopsis thaliana telomere repeat. Restriction fragments lacking highly repetitive DNA were identified and used as probes to map 14 of the 20 lambda clones. All of these markers mapped near the centromere on eight of the twelve tomato chromosomes. The exact centromere location of chromosomes 7 and 9 has recently been determined, and all ITR clones that localize to these two chromosomes map to the marker clusters known to contain the centromere. High-resolution mapping of one of these markers showed cosegregation of the telomere repeat with the marker cluster closest to the centromere in over 9000 meiotic products. We propose that the map location of interstitial telomere clones may reflect specific sequence interchanges between telomeric and centromeric regions and may provide an expedient means of localizing centromere positions.     

RFLP mapping of the genes controlling hybrid breakdown in rice (Oryza sativa L.)

 Complementary recessive genes hwd1 and hwd2 controlling hybrid breakdown (weakness of F₂ and later generations) were mapped in rice using RFLP markers. These genes produce a plant that is shorter and has fewer tillers than normal plants when the two loci have only one or no dominant allele at both loci. A cultivar with two dominant alleles at the hwd1 locus and a cultivar with two dominant alleles at the hwd2 locus were crossed with a double recessive tester line. Linkage analysis was carried out for each gene independently in two F₂ populations derived from these crosses. hwd1 was mapped on the distal region of rice genetic linkage map for chromosome 10, flanked by RFLP markers C701 and R2309 at a distance of 0.9 centiMorgans (cM) and 0.6 cM, respectively. hwd2 was mapped in the central region of rice genetic linkage map for chromosome 7, tightly linked with 4 RFLP markers without detectable recombination. The usefulness of RFLP mapping and map information for the genes controlling reproductive barriers are discussed in the context of breeding using diverse rice germplasm, especially gene introduction by marker-aided selection.     

Molecular-marker-facilitated studies of morphological traits in maize. II: Determination of QTLs for grain yield and yield components

Genetic factors controlling quantitative inheritance of grain yield and its components have not previously been investigated by using replicated lines of an elite maize (Zea mays L.) population. The present study was conducted to identify quantitative trait loci (QTLs) associated with grain yield and grain-yield components by using restriction fragment length polymorphism (RFLP) markers. A population of 150 random F₂₃ lines was derived from the single cross of inbreds Mo17 and H99, which are considered to belong to the Lancaster heterotic group. Trait values were measured in a replicated trial near Ames, Iowa, in 1989. QTLs were located on a linkage map constructed with one morphological and 103 RFLP loci. QTLs were found for grain yield and all yield components. Partial dominance to overdominance was the primary mode of gene action. Only one QTL, accounting for 35% of the phenotypic variation, was identified for grain yield. Two to six QTLs were identified for the other traits. Several regions with pleiotropic or linked effects on several of the yield components were detected.     

Molecular mapping of a major gene conferring resistance to maize mosaic virus

 The objective of this study was to determine the genetic basis of resistance to maize mosaic virus (MMV). Molecular markers were used to map resistance loci to MMV in a set of 91 maize (Zea mays L.) recombinant inbred lines (RILs), derived from the cross between Hi31 (a B68 conversion resistant to MMV) and Kil4 (a Thai inbred susceptible to MMV). The RILs were evaluated for MMV resistance in disease nurseries in Hawaii in the winter of 1993 and the summer of 1994. Twenty-eight highly susceptible RILs were chosen for gene mapping using the pooled-sampling approach. Initial evidence from the pooled DNA indicated that restriction fragment length polymorphism (RFLP) probes on chromosome 3 near the centromere were biased to the susceptible parent allele. Analysis of 91 RILs at 103 RFLP loci confirmed the presence of a major MMV resistance gene on chromosome 3. The resistant allele at this locus, previously named Mv1, is present in the resistant parent Hi31 and traces back to the Argentine parent used in conferring common rust resistance to B68. We conclude that resistance to MMV in B68 and Caribbean flints involves a major gene mv1 on chromosome 3 located between RFLP markers umc102 and php20508. Received: 12 June 1996 / Accepted: 5 July 1996