标记辅助导入中不同前景和背景选择方法的比较

标记辅助导入是分子遗传信息应用于动物育种的一个重要方面,其目的是在标记信息的辅助下将一个品种（供体）中的一个或多个优良基因导入另一个品种（受体）,同时还要尽可能地保持受体群体原有的遗传背景。标记辅助导入的过程包括3个阶段,第一阶段是杂交,即供体与受体杂交产生F1代个体,第二阶段是回交,即F1个体以及后续各个世代的后代个体重复地与受体回交,以使受体的遗传背景得到恢复,第三阶段是横交,即重复回交后得到的个体彼此问交配,以便获得供体基因的纯合个体,使该基因在群体中固定。在回交和横交阶段,都要对参与交配的个体进行选择。在选择中,要分别进行前景选择和背景选择,前景选择是对供体基因的选择,选择携带有供体基因个体参加配种,从而使该基因在回交过程中不会丢失,并在横交过程中能尽快固定,背景选择是对受体遗传背景的选择,选择那些含有受体基因组比例较高的个体参加配种,从而加快恢复受体遗传背景的速度。本研究通过计算机模拟对不同的前景选择方法和不同的背景选择方法进行了比较。前景选择方法包括对受体基因的直接选择（假设该基冈可以直接测定）、利用单个连锁标记的间接选择和利用两侧标记的间接选择3种,背景选择方法包括随机选择、基因组相似性选择、指数选择和标记辅助BLUP（MBLUP）选择4种。研究结果表明,对于前景选择来说,对供体基因的直接选择能保证该基因在回交的各个世代中保持一个稳定的频率（0．25）并在横交阶段迅速固定（2个世代）,用两侧标记的间接选择也能得到类似的结果,但如果仅利用单个连锁标记进行选择,则会导致供体基因的频率在回交阶段中有所下降,并在横交阶段不能被固定。对于背景选择来说,如果最终的目的是要完全恢复受体的遗传背景,基因组相似性选择或标记指数选择是最好的选择方法,它们可使受体的遗传背景在回交3个世代后就恢复到98％以上,而随机选择或MBLUP选择需要至少5个世代的回交才能达到这个水平。但如果最终的目的只是要恢复受体的某些优良性状,则MBLUP选择是值得推荐的方法,它可使影响这些性状的受体基因频率在回交3个世代后就达到99％以上,而且还能在整个基因导入过程中给这些性状带来最大的遗传进展。虽然用标记指数选择也有相似的结果,但与之相比,MBLUP的成本要低得多,更具有实际可行性。     

标记辅助导入的原理与应用

标记辅助导入是分子遗传信息应用于动物育种的一个重要方面, 其目的是在标记信息的辅助下将一个品种(供体)中的一个或多个优良基因导入另一个品种(受体), 同时还要尽可能地保持受体群体原有的遗传背景。在标记辅助导入过程中, 标记信息既可用于辅助前景选择, 即对目标基因携带者的选择, 以保证目标基因的正确导入, 也可用于辅助背景选择, 即对受体基因组的选择, 以加快受体遗传背景的恢复。本文介绍了标记辅助导入的原理和基本方法, 综述了目前已提出的不同前景选择和背景选择方法以及消除遗传累赘(与目标基因连锁的不利基因)的方法, 同时列举了标记辅助导入在动物上的一些成功应用。     

Stability over genetic backgrounds, generations and years of quantitative trait locus (QTLs) for organoleptic quality in tomato

The efficiency of marker-assisted backcross for the introgression of a quantitative trait locus (QTL) from a donor line into a recipient line depends on the stability of QTL expression. QTLs for six quality traits in tomato (fruit weight, firmness, locule number, soluble solid content, sugar content and titratable acidity) were studied in order to investigate their individual effect and their stability over years, generations and genetic backgrounds. Five chromosome regions carrying fruit quality QTLs were transferred following a marker-assisted backcross scheme from a cherry tomato line into three modern lines with larger fruits. Three sets of genotypes corresponding to three generations were compared: (1) an RIL population, which contained 50% of each parental genome, (2) three BC3S1 populations which segregated simultaneously for the five regions of interest but were almost fully homozygous for the recipient genome on the eight chromosomes carrying no QTL and (3) three sets of QTL-NILs (BC3S3 lines) which differed from the recipient line only in one of the five regions. QTL detection was performed in each generation, in each genetic background and during 2 successive years for QTL-NILs. About half of the QTLs detected in QTL-NILs were detected in both years. Eight of the ten QTLs detected in RILs were recovered in the QTL-NILs with the genetic background used for the initial QTL mapping experiment, with the exception of two QTLs for fruit firmness. Several new QTLs were detected. In the two other genetic backgrounds, the number of QTLs in common with the RILs was lower, but several new QTLs were also detected in advanced generations.     

Marker-assisted introgression using non-unique marker alleles I: selection on the presence of linked marker alleles

This paper investigates marker-assisted introgression of a major gene into an outbred line, where identification of the introgressed gene is incomplete because marker alleles are not unique to the base populations (the same marker allele can occur in both donor and recipient population). Those markers are used to identify the introgressed allele as well as the background genotype. The effect of using those markers, as if they were completely informative on the retention of the introgressed allele, was examined over five generations of backcrossing by using a single marker or a marker bracket for different starting frequencies of the marker alleles. Results were calculated by using both a deterministic approach, where selection is only for the desired allele, and by a stochastic approach, where selection is also on background genotype. When marker allele frequencies in donor and recipient population diverged from 1 and 0 (using a diallelic marker), the ability to retain the desired allele rapidly declined. Marker brackets performed notably better than single markers. If selection on background marker genotype was applied, the desired allele could be lost even more quickly than expected at random because the chance that the allele, which is common in the donor line, is present on the locus identifying the introgressed allele and is surrounded by alleles common in the recipient line on the background marker loci, will descend from the donor line (double recombination has taken place), is a lot smaller than the chance that this allele will stem from the recipient line (in which the allele occurs in low frequency). Marker brackets again performed better. Preselection against marker homozygotes (producing uninformative gametes) gave a slightly better retention of the introgressed allele.     

Combined detection and introgression of QTL in outbred populations

Background

Detecting a QTL is only the first step in genetic improvement programs. When a QTL with desirable characteristics is found, e.g. in a wild or unimproved population, it may be interesting to introgress the detected QTL into the commercial population. One approach to shorten the time needed for introgression is to combine both QTL identification and introgression, into a single step. This combines the strengths of fine mapping and backcrossing and paves the way for introgression of desirable but unknown QTL into recipient animal and plant lines.

Methods

The method consisting in combining QTL mapping and gene introgression has been extended from inbred to outbred populations in which QTL allele frequencies vary both in recipient and donor lines in different scenarios and for which polygenic effects are included in order to model background genes. The effectiveness of the combined QTL detection and introgression procedure was evaluated by simulation through four backcross generations.

Results

The allele substitution effect is underestimated when the favourable QTL allele is not fixed in the donor line. This underestimation is proportional to the frequency differences of the favourable QTL allele between the lines. In most scenarios, the estimates of the QTL location are unbiased and accurate. The retained donor chromosome segment and linkage drag are similar to expected values from other published studies.

Conclusions

In general, our results show that it is possible to combine QTL detection and introgression even in outbred species. Separating QTL mapping and introgression processes is often thought to be longer and more costly. However, using a combined process saves at least one generation. With respect to the linkage drag and obligatory drag, the results of the combined detection and introgression scheme are very similar to those of traditional introgression schemes.     

Introgression of a major QTL from an inferior into a superior population using genomic selection

Background

Selection schemes aiming at introgressing genetic material from a donor into a recipient line may be performed by backcross-breeding programs combined with selection to preserve the favourable characteristics of the donor population. This stochastic simulation study investigated whether genomic selection can be effective in preserving a major quantitative trait locus (QTL) allele from a donor line during the backcrossing phase.

Methods

In a simulation study, two fish populations were generated: a recipient line selected for a production trait and a donor line characterized by an enhanced level of disease resistance. Both traits were polygenic, but one major QTL affecting disease resistance was segregating only within the donor line. Backcrossing was combined with three types of selection (for total merit index) among the crossbred individuals: classical selection, genomic selection using genome-wide dense marker maps, and gene-assisted genomic selection. It was assumed that production could be observed directly on the selection candidates, while disease resistance had to be inferred from tested sibs of the selection candidates.

Results

Classical selection was inefficient in preserving the target QTL through the backcrossing phase. In contrast, genomic selection (without specific knowledge of the target QTL) was usually effective in preserving the target QTL, and had higher genetic response to selection, especially for disease resistance. Compared with pure genomic selection, gene-assisted selection had an advantage with respect to disease resistance (28–40% increase in genetic gain) and acted as an extra precaution against loss of the target QTL. However, for total merit index the advantage of gene-assisted genomic selection over genomic selection was lower (4–5% increase in genetic gain).

Conclusion

Substantial differences between introgression programs using classical and genomic selection were observed, and the former was generally inferior with respect to both genetic gain and the ability to preserve the target QTL. Combining genomic selection with gene-assisted selection for the target QTL acted as an extra precaution against loss of the target QTL and gave additional genetic gain for disease resistance. However, the effect on total merit index was limited.     

性状遗传力与QTL方差对标记辅助选择效果的影响

在采用动物模型标记辅助最佳线性无偏预测方法对个体育种值进行估计的基础上,模拟了在一个闭锁群体内连续对单个性状选择10个世代的情形,并系统地比较了性状遗传力和QTL方差对标记辅助选择所获得的遗传进展、QTL增效基因频率和群体近交系数变化的影响。结果表明：在对高遗传力和QTL方差较小的性状实施标记辅助选择时,可望获得更大的遗传进展;遗传力越高,QTL方差越大,则QTL增效基因频率的上升速度越快;遗传力较高时,群体近交系数上升的速度较为缓慢,而QTL方差对群体近交系数上升速度的影响则不甚明显。结合前人关于标记辅助选择相对效率的研究结果,可以认为：当选择性状的遗传力和QTL方差为中等水平时,标记辅助选择可望获得理想的效果。     

不同QTL增效基因初始频率下标记辅助选择的效果

采用随机模拟方法模拟了在一个闭锁群体内连续对单个性状选择10个世代的情形。在假定选择性状受一个位于常染色体上的QTL和多基因共同控制的情况下,采用动物模型标记辅助最佳线性无偏预测方法估计个体育种值并据此进行种畜的选留,并在此基础上系统地比较了QTL增效基因初始频率对标记辅助选择效果的影响。结果表明：当群体中QTL增效基因的初始频率较低时,选择所获得的QTL基因型值的进展会更大,标记辅助选择在单位时间内可获得较大的遗传进展;此时,尽管QTL增效基因在群体中固定所需的世代数会更长一些,但其频率上升的速度却更快。而QTL增效基因初始频率的高低对群体近交增量的影响不是很大。     

Evaluation of quantitative trait loci affecting intramuscular fat and reproductive traits in pigs using marker‐assisted introgression

We investigated the effects of previously identified quantitative trait loci (QTL) in an experimental backcross (BC) between Chinese Meishan pigs and commercial Duroc pigs. We performed marker‐assisted introgression of two QTL for intramuscular fat (IMF) content (IMF population) and three QTL for reproductive traits (reproduction population) from a donor Meishan pig into a recipient Duroc pig. At the fourth BC generation of the IMF population and third BC generation of the reproduction population, carrier animals were selected for the production of animals homozygous for the QTL. Our previous studies have shown that the presence of a Meishan allele on the IMF QTL is associated with low IMF values, and the Meishan allele on the reproductive QTL is associated with large litters. In this study, the presence of a Duroc allele at the IMF QTL on SSC9 resulted in a 0.27% increase in IMF (additive effect = 0.27 ± 0.08), whereas the presence of a Meishan allele at the IMF QTL on SSC7 resulted in a 0.34% increase in IMF (additive effect = ?0.34 ± 0.09). The presence of the Meishan allele at the IMF QTL on SSC7 thus had the opposite effect to our previous studies, that is, increased IMF. In the reproduction population, we observed no differences between the genotypes of the three QTL in regard to number of corpora lutea or litter size. Marker‐assisted introgression at these QTL is thus unlikely to result in an associated increase in litter size. These results show that it is possible to introgress alleles from other breeds into a selection population using molecular markers; any unexpected results might be associated with the genetic background.     

Genetic architecture of a selection response in Arabidopsis thaliana

Quantitative trait locus (QTL) mapping has become an established and effective method for studying the genetic architecture of complex traits. In this report, we use a QTL mapping approach in combination with data from a large selection experiment in Arabidopsis thaliana to explore a response to selection of experimental populations with differentiated genetic backgrounds. Experimental populations with genetic backgrounds derived from ecotypes Landsberg and Niederzenz were exposed to multiple generations of fertility and viability selection. This selection resulted in phenotypic shifts in a number of life-history and fitness-related characters including early development time, flowering time, dry biomass, longevity, and fruit production. Quantitative trait loci were mapped for these traits and their positions were compared to previously characterized allele frequency changes in the experimental populations (Ungerer et al. 2003). Quantitative trait locus positions largely colocalized with genomic regions under strong and consistent selection in populations with differentiated genetic backgrounds, suggesting that alleles for these traits were selected similarly in differentiated genetic backgrounds. However, one QTL region exhibited a more variable response; being positively selected on one genetic background but apparently neutral in another. This study demonstrates how QTL mapping approaches can be combined with map-based population genetic data to study how selection acts on standing genetic variation in populations.     

Marker-assisted selection to introgress rice QTLs controlling root traits into an Indian upland rice variety

A marker-assisted back-crossing (MABC) breeding programme was conducted to improve the root morphological traits, and thereby drought tolerance, of the Indian upland rice variety, Kalinga III. This variety, the recurrent parent in the MABC, had not previously been used for quantitative trait locus (QTL) mapping. The donor parent was Azucena, an upland japonica variety from Philippines. Five segments on different chromosomes were targeted for introgression; four segments carried QTLs for improved root morphological traits (root length and thickness) and the fifth carried a recessive QTL for aroma. Some selection was made at non-target regions for recurrent parent alleles. We describe the selection made in three backcross (BC) generations and two further crosses between BC₃ lines to pyramid (stack) all five target segments. Pyramids with four root QTLs were obtained in eight generations, completed in 6 years using 3,000 marker assays in a total of 323 lines. Twenty-two near-isogenic lines (NILs) were evaluated for root traits in five field experiments in Bangalore, India. The target segment on chromosome 9 (RM242-RM201) significantly increased root length under both irrigated and drought stress treatments, confirming that this root length QTL from Azucena functions in a novel genetic background. No significant effects on root length were found at the other four targets. Azucena alleles at the locus RM248 (below the target root QTL on chromosome 7) delayed flowering. Selection for the recurrent parent allele at this locus produced early-flowering NILs that were suited for upland environments in eastern India.     

Analysis of the Mesotelencephalic Dopamine System by Quantitative-Trait Locus Introgression

One of the significant factors that affect brain dopamine function is the activity of tyrosine hydroxylase (TH), the first and rate-limiting enzyme in catecholamine biosynthesis. For the analysis of the genetically determined role of dopamine function and TH in behavior and in the regulatory mechanisms of the mesotelencephalic dopamine system we devised a novel genetic strategy (Vadasz; Mouse Genome 88:16–18; 1990). We hypothesized that phenotypic introgression and recombinant fixation could ensure the transfer of Quantitative Trait Loci (QTL) from one strain onto the genetic background of another strain, and new, genetically very similar quasi-congenic strains could be created that would carry individual QTLs, or QTLs in various combinations. Here we summarize the construction of the first set of QTL Introgression strains, and present evidence that QTLs that are responsible for the continuous variation of mesencephalic tyrosine hydroxylase activity (TH/MES), have been transferred onto the C57BL/6By (B6) strain background from BALB/cJ (C) and CXBI (I) donor strains with high and low TH/MES, respectively. The QTL transfer was carried out in two directions by repeated backcross-intercross cycles with concomitant selection for the extreme high and low expressions of TH/MES in replicates, resulting in four QTL Introgression lines. Analysis of regional brain TH activities in the course of the QTL introgression indicated that (a) TH activity in B6.I lines showed quite limited heritability, (b) TH/MES was not highly correlated with striatal TH, and (c) the control of hypothalamic and olfactory tubercle TH activities was largely independent from that of TH/MES. Examination of the open-field (OF) behavior data demonstrated that TH activity did not correlate significantly with OF behavior. After 5 backcross-intercross cycles, TH/MES in each replicate line was still significantly different from that of the B6 background strain. A genomewide scanning of microsatellite markers in the QTL introgression lines demonstrated that about 96% of the markers were of background (B6) type. These results indicate the successful transfer of TH/MES QTLs. After the QTL transfer phase of the experiment altogether more than 100 new RQI strains were initiated in the QTL Introgression lines by strict brother × sister mating. After fixing the introgressed QTLs, ten of the inbred RQI strains were tested for TH/MES. The results showed that in one of the new RQI strains TH/MES was restored to a level that is characteristic to the C donor strain, while TH/MES values in some other strains were between those of the background and donor strains, confirming our hypothesis that phenotypic introgression and recombinant fixation can ensure a virtually complete transfer of QTLs. We conclude from this study that complex, continuously distributed neural traits can successfully be subjected to QTL introgression, and the results raise the possibility that the RQI method can be efficiently applied for gene mapping of complex neural and behavioral traits even if their phenotypic expression is sensitive to confounding developmental and environmental variations, genetic interactions, and genotype-environment interactions.     

Marker-assisted introgression of Trypanotolerance QTL in mice

A marker-assisted introgression (MAI) experiment was conducted to use genetic markers to transfer each of the three trypanotolerance QTL from a donor mouse strain, C57BL/6, into a recipient mouse strain, A/J. We used a backcross strategy that consisted of selecting two lines, each carrying two of the donor QTL alleles through the backcross (BC) phase. At the fourth BC generation, single-carrier animals were selected for the production of homozygous animal in the intercross phase. The QTL regions (QTLR) were located on chromosomes MMU1, MMU5, and MMU17. Groups of mice with different genotypes and the parental lines were subjected to a challenge with Trypanosoma congolense. The results show that trypanotolerance QTL was successfully moved into the recipient background genotype, yielding a longer survival time. The mean estimated survival time was 57.9, 49.5, and 46.8 days for groups of mice carrying the donor QTL on MMU1, MMU5, and MMU17 on A/J background. The mean estimated survival time was 29.7 days for the susceptible A/J line and 68.8 days for the resistant C57BL/6 line. The estimated QTLR effects are close to 30% smaller than those in the original mapping population which was likely caused by the difference in the background on which the effects of QTLR are tested. This is the first report of successful marker-assisted introgression of QTL in animals. It is experimental proof of the use of genetic markers for marker-assisted introgression in animal breeding.Institut National des Recherches Agricoles du Bénin, 01 BP 884 COTONOU, République du Bénin     

Genome‐wide analysis of allele frequency change in sunflower crop–wild hybrid populations evolving under natural conditions

Crop‐wild hybridization occurs in numerous plant species and could alter the genetic structure and evolutionary dynamics of wild populations. Studying crop‐derived alleles in wild populations is also relevant to assessing/mitigating the risks associated with transgene escape. To date, crop‐wild hybridization has generally been examined via short‐term studies, typically within a single generation, focusing on few traits or genetic markers. Little is known about patterns of selection on crop‐derived alleles over multiple generations, particularly at a genome‐wide scale. Here, we documented patterns of natural selection in an experimental crop × wild sunflower population that was allowed to evolve under natural conditions for two generations at two locations. Allele frequencies at a genome‐wide collection of SNPs were tracked across generations, and a common garden experiment was conducted to compare trait means between generations. These data allowed us to identify instances of selection on crop‐derived alleles/traits and, in concert with QTL mapping results, test for congruence between our genotypic and phenotypic results. We found that natural selection overwhelmingly favours wild alleles and phenotypes. However, crop alleles in certain genomic regions can be favoured, and these changes often occurred in parallel across locations. We did not, however, consistently observe close agreement between our genotypic and phenotypic results. For example, when a trait evolved towards the wild phenotype, wild QTL alleles associated with that trait did not consistently increase in frequency. We discuss these results in the context of crop allele introgression into wild populations and implications for the management of GM crops.     

The construction of a substitution library of recombinant backcross lines in Brassica oleracea for the precision mapping of quantitative trait loci.

The currently available methods for locating quantitative trait loci (QTLs) and measuring their effects in segregating populations lack precision unless individual QTLs have very high heritabilities. The use of recombinant backcross lines containing short regions of donor chromosome introgressed into a constant recipient background permits QTLs to be located with greater precision. The present paper describes the use of molecular markers to introgress defined short regions of chromosome from a donor doubled haploid calabrese line of Brassica oleracea (var. italica) into a recipient short generation variety (Brassica oleracea var. alboglabra). We demonstrate that in just two or three generations of backcrossing, combined with selection for mapped molecular markers, the generation of a library of recombinant backcross lines is feasible. The possible use and refinement of these lines are discussed. Key words : backcrossing, Brassica oleracea, introgression, molecular markers, near-isogenic lines, QTL mapping, recombinant backcross lines, substitution lines.     

Simultaneous improvement and genetic dissection of grain yield and its related traits in a backbone parent of hybrid rice (Oryza sativa L.) using selective introgression

Three populations with a total of 125 BC2F3:4 introgression lines (ILs) selected for high yields from three BC2F2 populations were used for genetic dissection of rice yield and its related traits. The progeny testing in replicated phenotyping across two environments and genotyping with 140 polymorphic simple sequence repeat markers allowed the identification of 21 promising ILs that had significantly higher yields than the recurrent parent Shuhui527 (SH527). A total of 94 quantitative trait loci (QTL) were identified using the selective introgression method based on Chi-squared (χ ²) and multi-locus probability tests and the RSTEP-LRT method based on stepwise regression. These QTL were mostly mapped to 12 clusters on seven rice chromosomes. Several important properties of the QTL affecting grain yield (GY) and its related traits were revealed. The first one was the presence of strong and frequent non-random associations between or among QTL that affect low-heritability traits (GY and spikelet number per panicle, SN) in the ILs with high trait values. Second, beneficial alleles at 88.9 % GY and 75 % SN QTL for increased productivity were from the donors, suggesting that direct phenotypic selection for high yield in our introgression breeding program was a powerful way to transfer beneficial alleles at many loci from the donors into SH527. Third, most QTL were in clusters with large effects on multiple traits, which should be the focal points in further investigations and marker-assisted selection in rice. The majority of the QTL identified were expressed only in one of the environments, suggesting that differential expression of QTL in different environments is the primary genetic basis of genotype × environment interaction. Finally, a large variation in both the direction and magnitude of QTL effects was detected for different donor alleles at seven QTL in the same genetic background and environments. This finding suggests the possible presence of functional diversity among the donor alleles at these loci. The promising ILs and QTL identified provide valuable materials and genetic information for further improving the yield potential of SH527, which is a backbone restorer of hybrid rice in China.     

Genomic contributions in livestock gene introgression programmes

The composition of the genome after introgression of a marker gene from a donor to a recipient breed was studied using analytical and simulation methods. Theoretical predictions of proportional genomic contributions, including donor linkage drag, from ancestors used at each generation of crossing after an introgression programme agreed closely with simulated results. The obligate drag, the donor genome surrounding the target locus that cannot be removed by subsequent selection, was also studied. It was shown that the number of backcross generations and the length of the chromosome affected proportional genomic contributions to the carrier chromosomes. Population structure had no significant effect on ancestral contributions and linkage drag but it did have an effect on the obligate drag whereby larger offspring groups resulted in smaller obligate drag. The implications for an introgression programme of the number of backcross generations, the population structure and the carrier chromosome length are discussed. The equations derived describing contributions to the genome from individuals from a given generation provide a framework to predict the genomic composition of a population after the introgression of a favourable donor allele. These ancestral contributions can be assigned a value and therefore allow the prediction of genetic lag.     

Marker-Assisted Introgression of Quantitative Trait Loci

The use of molecular markers for the introgression of one or several superior QTL alleles into a recipient line is investigated using analytic and simulation results. The positions of the markers devoted to the control of the genotype at the QTLs in a ``foreground selection' step are optimized given the confidence interval of the QTL position. Results demonstrate that using at least three markers per QTL allows a good control over several generations. Population sizes that should be recommended for various numbers of QTLs are calculated and are used to determine the limit in the number of QTLs that can be monitored simultaneously. If ``background selection' devoted to accelerate the return to the recipient parent genotype outside the QTL regions is applied, the positions of the markers devoted to the control of the QTLs have to be reconsidered. When several QTLs are monitored simultaneously, background selection among the limited number of individuals resulting from the foreground selection step accelerates the increase in genomic similarity with the recipient parent, with only limited costs. Background selection is even more efficient in a pyramidal backcross program where QTLs are first monitored one by one.     

Optimized breeding strategies for multiple trait integration: I. Minimizing linkage drag in single event introgression

From a breeding standpoint, multiple trait integration (MTI) is a four-step process of converting an elite variety/hybrid for value-added traits (e.g. transgenic events) using backcross breeding, ultimately regaining the performance attributes of the target hybrid along with reliable expression of the value-added traits. In the light of the overarching goal of recovering equivalent performance in the finished conversion, this study focuses on the first step of MTI, single event introgression, exploring the feasibility of marker-aided backcross conversion of a target maize hybrid for 15 transgenic events, incorporating eight events into the female hybrid parent and seven into the male parent. Single event introgression is conducted in parallel streams to convert the recurrent parent (RP) for individual events, with the primary objective of minimizing residual non-recurrent parent (NRP) germplasm, especially in the chromosomal proximity to the event (i.e. linkage drag). In keeping with a defined lower limit of 96.66 % overall RP germplasm recovery (i.e. ≤120 cM NRP germplasm given a genome size of 1,788 cM), a breeding goal for each of the 15 single event conversions was developed: <8 cM of residual NRP germplasm across the genome with ~1 cM in the 20 cM region flanking the event. Using computer simulation, we aimed to identify optimal breeding strategies for single event introgression to achieve this breeding goal, measuring efficiency in terms of number of backcross generations required, marker data points needed, and total population size across generations. Various selection schemes classified as three-stage, modified two-stage, and combined selection conducted from BC1 through BC3, BC4, or BC5 were compared. The breeding goal was achieved with a selection scheme involving five generations of marker-aided backcrossing, with BC1 through BC3 selected for the event of interest and minimal linkage drag at population size of 600, and BC4 and BC5 selected for the event of interest and recovery of the RP germplasm across the genome at population size of 400, with selection intensity of 0.01 for all generations. In addition, strategies for choice of donor parent to facilitate conversion efficiency and quality were evaluated. Two essential criteria for choosing an optimal donor parent for a given RP were established: introgression history showing reduction of linkage drag to ~1 cM in the 20 cM region flanking the event and genetic similarity between the RP and potential donor parents. Computer simulation demonstrated that single event conversions with <8 cM residual NRP germplasm can be accomplished by BC5 with no genetic similarity, by BC4 with 30 % genetic similarity, and by BC3 with 86 % genetic similarity using previously converted RPs as event donors. This study indicates that MTI to produce a ‘quality’ 15-event-stacked hybrid conversion is achievable. Furthermore, it lays the groundwork for a comprehensive approach to MTI by outlining a pathway to produce appropriate starting materials with which to proceed with event pyramiding and trait fixation before version testing.     

Factors influencing the efficiency of a marker-assisted introgression programme in Merino sheep

This study investigated a marker-assisted introgression programme in Australian Merino sheep. The goal was to introgress an allele with a large negative effect on fibre diameter into a Merino flock possessing medium average fibre diameter. The influence of two factors was explored: the strategy used to select animals from the purebred and backcross line for backcrossing purposes and the use of selection on background markers to accelerate the return to the purebred line''s genome. The results were compared to introgression based on EBVs only. Introgression using EBVs only produced almost the same response in the dollar index as marker-based introgression methods. However, this study did not account for some of the costs associated with implementing the programmes, including the costs of phenotyping and genotyping. Given that the cost of measuring fibre diameter is low, it was concluded that introgression on EBVs only would be the preferred method since the marginal profit of marker-assisted introgression would not be large enough to cover the additional cost of genotyping. In marker-assisted introgression, reciprocal crossing of male and female selection candidates from the backcross and the purebred line was the most advantageous strategy from a practical and profit point of view. Selection for background markers was less profitable in this study than recovering the donor genome by selection on phenotype.