Potential of marker-assisted selection in hemp genetic improvement

Summary The development and applications of molecular markers to hemp breeding are recent, dating back only to the mid-1990s. The main achievements in this field are reviewed. The analysis of Cannabis germplasm by RAPD, AFLP and microsatellites is discussed, with its consequence for the still debated species concept in Cannabis. DNA-based markers have also been exploited in the field of forensic science, in an attempt to discriminate licit from illicit crop. The main applications of the molecular markers to the breeding, however, have been achieved with the development of markers closely linked to the male sex and to some of the most relevant chemotypes. Active research is carried out by several groups in the field of identification and characterization of the genes involved in fiber quality and quantity, and in the determination of monoecy, another very important target of hemp breeding. Besides, markers associated to new, potentially useful chemotypes are being developed, for the marker-assisted breeding of pharmaceutical Cannabis.     

Identification of QTLs for rice grain size and shape of Iranian cultivars using SSR markers

Grain size is a main component of rice appearance quality. In this study, we performed the SSR mapping of quantitative trait loci (QTLs) controlling grain size (grain length and breadth) and shape (length/breadth ratio) using an F₂ population of a cross between two Iranian cultivars, Domsephid and Gerdeh, comprising of 192 individuals. A linkage map with 88 markers was constructed, which covered 1367.9 cM of the rice genome with an average distance of 18 cM between markers. Interval mapping procedure was used to identify the QTLs controlling three grain traits, and QTLs detected were further confirmed using composite interval mapping. A total of 11 intervals carrying 18 QTLs for three traits were identifed, that included five QTLs for grain length, seven QTLs for grain breadth, and six QTLs for grain shape. A major QTL for grain length was detected on chromosome 3, that explained 19.3% of the phenotypic variation. Two major QTLs for grain breadth were mapped on chromosomes 3 and 8, which explained 34.1% and 20% of the phenotypic variation, respectively. Another two major QTLs were identified for grain shape on chromosomes 3 and 8, which accounted for 27.1% and 20.5% of the phenotypic variance, respectively. The two QTLs that were mapped for grain shape coincided with the major QTLs detected for grain length and grain breadth. Intrestingly, gs2 QTL specific to grain shape was detected on chromosome 2 that explained 15% of the phenotypic variation.     

Use of molecular markers to accelerate the breeding of common bean lines resistant to rust and anthracnose

The main goal of this work was to introduce resistance genes for rust, caused by Uromyces appendiculatus, and anthracnose, caused by Colletotrichum lindemuthianum, in an adapted common bean cultivar through marker-assisted backcrossing. DNA fingerprinting was used to select plants genetically closer to the recurrent parent which were also resistant to rust and to race 89 of C. lindemuthianum. DNA samples extracted from the resistant parent (cv. Ouro Negro), the recurrent parent (cv. Rudá), and from BC₁, BC₂ and BC₃ resistant plants were amplified by the RAPD technique. The relative genetic distances in relation to the recurrent parent varied between 9 and 59% for BC₁, 7 and 33% for BC₂, and 0 and 7% for BC₃ resistant plants. After only three backcrosses, five lines resistant to rust and anthracnose with, approximately, 0% genetic distance in relation to the recurrent parent were obtained. These lines underwent field yield tests in two consecutive growing seasons and three of them presented a good yield performance, surpassing in that sense their parents and most of the reference cultivars tested.     

Two major genes for kernel weight in rye

Previously, kernel weight in rye was considered mostly as a quantitatively inherited trait. F₁ plants from parents showing a great difference in kernel weight were selfed and the segregating F₂ population was analysed with molecular markers. Two simple sequence repeats markers, one on chromosome 5 and the other on chromosome 7, were found which allowed the genetic analysis of two major genes acting in a complementary way. The analysis of major genes is an advantage in breeding for this important economic trait. In addition it can provide more insight into the genetic structure of this character. This is, in turn, also a prerequisite for future investigations of this trait at the molecular level.     

Homoeological relationships between the f chromosome of Brassica rapa and the e chromosome of Brassica oleracea

Eight plants of the putative double monosomic addition line (DMAL, 2n= 20) were developed by crossing a monosomic chromosome addition line of radish [f(A)‐type monosomic addition line (MAL) (2n= 19)] carrying the f chromosome of Brassica rapa (2n= 20, AA) with another [e(C)‐type MAL (2n= 19)] having the echromosome of Brassica oleracea (2n= 18, CC). The homoeological relationships between the two alien chromosomes were investigated by morphological, cytogenetic and random amplified polymorphic DNA (RAPD) analysis. Seventeen morphological traits that were not present in the radish cv. ‘Shogoin’ were observed in both MALs and these traits were substantially exhibited in DMAL plants. At the first metaphase of pollen mother cells (PMCs), the two parental MALs showed a chromosome configuration of 9II +1I, demonstrating impossibility of recombination between the R and the added chromosomes. The DMALs formed 10II in approximately 73% of PMCs, with one bivalent showing loose pairing between two chromosomes differing in size. In an attempt to identify the two MALs by RAPD‐specific markers using 26 selected random primers, 13 and 20 bands were specific for the f(A)‐type and the e(C)‐type MALs, respectively; 12 bands were common to both MALs (26.7%). In conclusion, the f chromosome of B. rapa is homoeologous to the e chromosome of B. oleracea. The genetic domain (genes) for 17 morphological traits are linked to each homoeologous chromosome bearing 27% of the corresponding RAPD markers.     

Progress in mapping agronomic genes in apple (The European Apple Genome Mapping Project)

Summary The progress of the European Apple Genome Mapping Project is described. Populations segregating for a range of agronomic genes have been established in six European countries. The need for robust methods of analysis has been identified, especially with regard to the development of molecular markers. Isozyme systems, RAPDs, RFLPs and amplified genes are being used to construct a reference genetic linkage map. Standardisation and precise definition of both genotypic and phenotypic measurements has been recognised as being essential for future exploitation of genetic markers in apple breeding. Phenotypic measurements are being replicated in different geographical locations over several years. Statistical and genetic analyses are aimed at defining components of genetic variation which account for ‘genes’, as defined by apple breeders. A relational database is being constructed which will combine disparate sources of data relating to the genetics of apple. Comparative mapping has been identified as an efficient means of expanding genetic knowledge within and between Rosaceae genomes.     

Chromosomal location of genes for resistance to powdery mildew in common wheat (Triticum aestivum L. em Thell.). 9. Gene MlZec1 from the Triticum dicoccoides-derived wheat line Zecoi-1

The Triticum dicoccoides-derived wheat line Zecoi-1 provides effective protection against powdery mildew. F₃ segregation analysis of Chinese Spring × Zecoi-1 hybrids showed that resistance in line Zecoi-1 is controlled by a single dominant gene. Amplified fragment length polymorphism (AFLP) analysis of bulked segregants from F₃s showing the homozygous resistant and susceptible phenotypes identified eight markers, of which four were associated with the resistance allele in repulsion phase. Following the assignment of these four repulsion phase AFLP markers to wheat chromosome 2B with the aid of Chinese Spring nulli-tetrasomic lines, they were physically mapped in the terminal breakpoint interval 0.89 (2BL-6)–1.00 (telomere) of chromosome 2BL. Genetic and physical mapping of simple sequence repeat markers from the distal half of chromosome 2BL located the wild emmer-derived powdery mildew resistance gene distal of breakpoint 0.89 in deletion line 2BL-6. Based on disease response patterns, genomic origin and chromosomal location the resistance gene in Zecoi-1 is temporarily designated MlZec1.     

Genetic analysis of a CSR10 (indica) × Taraori Basmati F3 population segregating for salt tolerance using ISSR markers

Segregation for salinity tolerance and ISSR markers based molecular polymorphism were investigated in a F₃ plant population raised via single-seed descent method from a cross between salt-tolerant indica rice variety CSR10 and salt-susceptible premium traditional Basmati rice variety Taraori Basmati HBC19. A total of 130 F₃plants were evaluated individually for salinity tolerance on 1–9 scale on the basis of seedling growth parameters; the average score ranged between 1.7 to 8.3. Frequency distribution curve obtained using the salinity tolerance data of F₃ population and a chi-square analysis, showed a good fit to a normal distribution. Eleven plants each in the category of salt-tolerant and salt-susceptible were selected from the segregating F₃ population for ISSR marker analysis. A total of 149 bands (4–11 bands per primer) ranging from 200 to 3530 bp were scored for the two rice varieties and the selected CSR10 × HBC19 segregating F₃ plants using 26 ISSR primers. Of these, 89 were monomorphic and 60 were polymorphic. Of the 60 polymorphic bands,36 and 20 bands were specific to CSR10 andHBC19 respectively. The remaining four bands were amplified using UBC primers 810,848, 853 and 886 and present in only some of the CSR10 × HBC19 F₃ plants. Notably, ISSR primers with dinucleotide repeat motif and 5'-anchored end amplified more number of bands (7.0 bands/primer) compared to3'-anchored dinucleotide primers (5.4bands/primer), but 3'-anchored dinucleotide primers revealed higher level of polymorphism (2.6 polymorphic bands/primer) compared to 5'-anchoreddinucleotide primers (1.43 polymorphic bands/ primer). While distribution of majority of the polymorphic bands were more or less in the expected ratios in salt-tolerant and/or salt-sensitive F₃segregating plants, but some of the bands amplified using UBC ISSR primers 823, 825,826, 849, 853, 864, 866 and 884 showed highly skewed distribution. Such polymorphic bands stand greater chances of having a linkage with the genes/ QTLs for salinity tolerance and shall be the target for further studies. This revised version was published online in July 2006 with corrections to the Cover Date.     

Use of multiparental inbred populations to determine allelic relationships of molecular markers

The assessment of polymorphism exhibited by molecular markers is an arduous but essential task that facilitates the use of molecular tools by breeders and geneticists. For that purpose, the value of a wheat composite population was assessed for characterizing the diversity of restriction fragment length polymorphism (RFLP) markers developed by INRA-Génoblé. The polymorphism of 13 genomic probes was measured over a set of 80 inbred lines randomly extracted by single-seed descent from a composite-cross of 16 wheat lines. The 13 probéenzyme combinations revealed 27 loci with codominant polymorphism. As many bands were so far unmapped, the segregational analysis of the progenies appeared very suitable for complex patterns, both in determining allelic relationships and in revealing linkage between loci. Allelic diversity, band sizes and chromosomal location assessed from nullisomic-tetrasomic lines are given for the 27 loci.     

Sex-linked SSR markers in hemp

Hemp is a dioecious plant with sex chromosomes X and Y, the male sex being heterogametic. The quality of the fibre depends on the sex type. The sex chromosomes can be characterized by molecular markers. In this report, sex‐linked simple sequence repeat (SSR) markers are described. One SSR marker was polymorphic in both the populations derived from single crosses, two other markers in but one of the two populations. Three alleles were detected for two SSR markers indicating polymorphism not only between X and Y, but also between different X chromosomes. In addition, several sex‐linked RAPD markers were detected in one population. Recombination within the sex chromosomes was observed for nearly all markers.