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2. Molecular Genetics 2.1. Marker-assisted selection

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Marker-assisted selection has been exploited to identify papaya genes that are associated with disease resistance or fruit quality traits. Research has focused on identifying markers for PRSV in Vasconcellea species (Drew et al., 1998; Dillon et al., 2006a; O’Brien and Drew, 2010; Kanchana-udomkan et al., 2018) and sweetness markers for improved fruit (Nantawan et al., 2018).

A protein kinase marker linked to the prsv-1 marker enabled development and use of sequence-characterized amplified region (SCAR) and cleavage amplified polymorphic sequence (CAPS) markers to track breeding of the resistance gene from V. pubescens into V. parviflora into papaya (Dillon et al., 2005a,b, 2006a,b; O’Brien and Drew, 2009; Kanchana-udomkan et al., 2018). The resistance gene was to be transformed into papaya; however, due to controversies over engineered plants, the research was never finalized (Razali, 2013; Razean Haireen and Drew, 2016; R. Razali, Malaysia, 2018, personal communication).

Productivity and fruit quality traits were evaluated in c.30 Australian cultivars (Nantawan et al., 2016). Sweetness-associated genes for marker development were identified through differential expression analyses of two different papaya cultivars, ‘Sunrise’ and ‘RB2’ (Nantawan et al., 2018). Differences in expression of sucrose phosphate synthase and invertase genes may be important, although studies are still in the early stages.

There has been ongoing interest in identifying the PRSV resistance gene in V. pubescens using marker-assisted selection. After genetic mapping of hybrids with randomly amplified DNA fingerprint (RAF) markers (Waldron et al., 2002; Peace et al., 2003; Ramage et al., 2003) using a large F2 population of 268 individuals, several F1 plants were sib-mated from among F1 plants involving V. pubescens (PRSV resistant) × V. parviflora (PRSV susceptible) progeny. The F2 plants were inoculated with PRSV, and a resistance locus, prsv-1, was identified on linkage group 7 in the V. pubescens map (Drew et al., 1998; Dillon et al., 2005a, 2005b, 2006b; C. O’Brien, R. Drew, unpublished results). The RAF marker, OPK41R, was converted to a SCAR marker and segregated perfectly with PRSV resistance in the F2 plants, was not amplified in papaya DNA and was converted to a CAPS marker, PsiIk4, after digestion with PsiI (Dillon et al., 2006b). Mapping to within 2 cM of the resistance locus, the CAPS marker was used to identify 99% of the resistant hybrids, making it invaluable for identifying resistance in V. pubescens hybrids with papaya. The CAPS marker is a useful tool for marker-assisted breeding programmes aimed at introgressing PRSV resistance genes from V. pubescens into commercially important papaya cultivars (Dillon et al., 2006b).

Since V. pubescens × papaya hybridizations resulted in PRSV-resistant (Manshardt and Wenslaff, 1989b) but sterile F1 hybrids (O’Brien and Drew, 2009), bridge-crosses of V. pubescens × V. parviflora were made to transfer the PRSV resistance gene into hybrids with larger proportions of the V. parviflora genome (O’Brien and Drew, 2009). The SCAR and CAPS markers (Dillon et al., 2006a, b) were used to trace the V. pubescens resistance gene in backcrosses to V. parviflora up to BC4 (Kanchana-udomkan et al., 2018). The markers were also used to identify and clone the V. pubescens gene containing the prsv-1 locus to transform directly into papaya to determine if transgenic plants would be virus resistant (Razali, 2013). Line #113 was crossed with papaya in different locations to produce F1s that were backcrossed with papaya. Eighteen hybrids with papaya were obtained that flowered and were crossed with papaya for the first backcross; however, there was no seed set. Groups in India and Hawaii used embryo rescue to recover resistance gene-bearing F1s with papaya, but subsequent crosses with papaya failed to produce seeds. Flowers on two F1s in Hawaii were pink, resembling the V. parviflora parent. One plant was female with single flowers per node and the other, hermaphrodite, with panicles of male-like flowers that bore green stigmas. The V. parviflora and BC4 plants grow vigorously during the spring, summer and autumn when it is warm, but were dormant in winter and dropped their leaves. The F1s with papaya remain leafy year round but grow very slowly and only flower in mid-winter or early spring. About 35 of the 66 embryo-rescued F1 lines contained the resistance gene (M. Fitch, X. He, unpublished results).

A review of marker-assisted selection (Kanchana-udomkan et al., 2016b) contains extensive citations and includes early observations by Storey (1938). Certain topics pertinent to papaya improvement, e.g. the identity of the V. pubescens and V. quercifolia genes that confer resistance to PRSV, are described in recent reviews (Kanchana-udomkan et al., 2016, 2018).

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