is rr TT WW; red is rr tt WW; and white with pink tip is rr tt ww. Seed size is controlled by several genes, including l (long) and s (short seed) genes, with s epistatic to l, and long recessive to medium or short. The phenotypes are LL SS for medium, ll SS for long, and LL ss or ll ss for short seed. The sources for genotypes are ll SS from ‘Peerless’, LL SS from ‘Klondike’, and LL ss from ‘Baby Delight’. A third gene, ts, produces tomato seed size, which is smaller than short seed. Also, tiny seed (Ti, dominant to short seed) from ‘Sweet Princess’ produces seeds that are between short and tomato seed sizes. Five genes for seed protein composition (Spr-1, Spr-2, Spr-3, Spr-4 and Spr-5) are included in the watermelon gene list.
Four linkage groups of 13 isozyme genes were reported by Navot and Zamir (1986). Since then, the Citrullus genome has been sequenced using the Chinese breeding line 97103 (Guo et al., 2013), and later using ‘Charleston Gray’.
With the advent of next-generation sequencing technologies, genomes of some of the major cucurbits have been sequenced, creating new sets of resources for these crops. Genomes of watermelon, melon, cucumber, squash (C. pepoand C. moschata), pumpkin (C. maxima), and bottle gourd (L. siceraria) are now available, and the list will continue to grow as sequencing costs decrease (see Table 3.1). The information is available on the Cucurbit Genomics Database maintained by J. Fei at Boyce Thompson Institute (Cornell University, Ithaca, New York).
Cucumber, the first cucurbit genome to be sequenced (2009), enabled scientists, including plant breeders, to identify novel biosynthetic pathways and to develop molecular markers. With each cucurbit species genome that has been sequenced, we have obtained a greater understanding of the evolution of the Cucurbitaceae as well as the function of genes shared across species. The high synteny between genetic sequences of cucumber and melon showed the chromosome fusions that led to cucumber from melon. The bottle gourd genome, one of the most recent species sequenced, brought new evidence that ancient cucurbits had 12 chromosomes. While most major cucurbit crops have had whole-genome sequencing completed, many minor cucurbits have had partial genome sequencing performed. Through these studies, novel and species-specific genes have been identified for bitter gourd (Behera et al., 2016), wax gourd (Jiang et al., 2013), wild Cucumis species (Ling et al., 2017) and other cucurbits, including the creeping perennial, Gynostemma pentaphyllum (Chen et al., 2016).
These resources have been leveraged for dozens of studies on transcriptome analyses of developing fruit, disease resistance or susceptibility, growth under abiotic stress conditions, and many other stages of plant growth in cultivated and wild cucurbits. Many transcriptome studies to date in cucurbits have focused on cucumber because of its economic importance (Ando et al., 2012; Kong et al., 2015; Wyatt et al., 2016; Li et al., 2017; Sun et al., 2017). While few cucurbit transcriptome studies have resulted in the identification of a single gene controlling a specific trait, these large-scale transcriptome studies provide valuable information on genetic pathways, patterns of expression, and candidates for breeding and genetic transformation technologies.
HISTORICAL BREEDING TECHNIQUES
The nature of DNA in cucurbits has been under study for more than three decades. Bhave et al. (1986) analysed the distribution of repeat and single copy DNA sequences in smooth and angled luffa, wax gourd and ivy gourd. Around the same time, Ganal and Hemleben (1986) compared restriction enzyme maps of ribosomal DNA repeats in cucumber, melon, and two squash species, C. maxima and C. pepo. An early and continuing objective for studying DNA and RNA in cucurbits has been to clarify phylogenetic relationships within the family. Analysis of variation of chloroplast DNA has revealed species relationships in Cucurbita (Wilson et al., 1992) and Cucumis (Perl-Treves and Galun, 1985), and researchers continue to use DNA-based information systems to detect origins and evaluate relationships in the Cucurbitaceae (Schaefer et al., 2009; Chomicki and Renner, 2014).
Speed breeding is a system that makes use of new technology to develop cultivars in less time. Some of those technologies that have been applied to speed breeding of cucurbits are as follows. Greenhouses can be supplied with lights that permit faster growth and flowering in the winter season. In addition, temperature control permits rapid growth, usually with the objective of 32°C day and 21°C night. Field trials can be run faster by using many locations and few years. Speed breeding usually builds on previous technologies. Some of those include winter nurseries, optimum size of roots in greenhouse pots or bags, and the use of growth regulators to increase the speed of making crosses and self-pollinations.
Much of the early progress made towards understanding the genetic code of cucurbits and other plants was due to the use of restriction enzymes, also called endonucleases. An endonuclease cleaves DNA at a particular location based on the enzyme’s recognition of a certain sequence of nucleotides. There are many different endonucleases, each cutting DNA at different places, and creating DNA fragments of various lengths. Endonucleases allowed for DNA mapping, which has been going on for cucumber, melon and squash for the past 30 years. For example, Gounaris et al. (1990) used endonucleases to resolve the genetic map of chloroplast DNA in C. pepo, and more recently, endonucleases were used to generate a genetic map of zucchini (C. pepo) using genotyping by sequencing (GBS) (Montero-Pau et al., 2017).
Endonucleases also enable scientists to create recombinant DNA, which results when cleaved DNA strands from two different DNA molecules are enzymatically joined. If one of these molecules is a bacterial plasmid (an independent, self-replicating ring of DNA in the bacterial cell), then DNA from the other molecule can be incorporated into the plasmid. The plasmid containing the recombinant DNA can be transferred to embryonic plant cells, which, in turn, pass the genetic material to new cells.
When the transgenic vector is a bacteriophage (a bacteria-infecting virus) instead of a plasmid, the recombinant DNA is passed from virus to bacterium (where it is incorporated into the bacterial DNA), and then from bacterium to plant. Bacteria containing incorporated viral DNA possess a degree of immunity to attacks by the same phage because the incorporated DNA directs synthesis of a phage repressor molecule that regulates the expression of the viral DNA.
Speed breeding has advanced as new technologies have emerged. Doubled haploid production has made it faster to produce inbred lines, although it is sometimes difficult
