Abstract
The seed oil of Jatropha curcas is considered a potential bioenergy source that could replace fossil fuels. However, the seed yield of Jatropha is low and has yet to be improved. We previously reported that exogenous cytokinin treatment increased the seed yield of Jatropha. Cytokinin levels are directly regulated by isopentenyl transferase (IPT), cytochrome P450 monooxygenase, family 735, subfamily A (CYP735A), and cytokinin oxidase/dehydrogenase (CKX). In this study, we cloned six IPT genes, one JcCYP735A gene, and seven JcCKX genes. The expression patterns of these 14 genes in various organs were determined using real-time quantitative PCR. JcIPT1 was primarily expressed in roots and seeds, JcIPT2 was expressed in roots, apical meristems, and mature leaves, JcIPT3 was expressed in stems and mature leaves, JcIPT5 was expressed in roots and mature leaves, JcIPT6 was expressed in seeds at 10 days after pollination, and JcIPT9 was expressed in mature leaves. JcCYP735A was mainly expressed in roots, flower buds, and seeds. The seven JcCKX genes also showed different expression patterns in different organs of Jatropha. In addition, CK levels were detected in flower buds and seeds at different stages of development. The concentration of N6-(Δ2-isopentenyl)-adenine (iP), iP-riboside, and trans-zeatin (tZ) increased with flower development, and the concentration of iP decreased with seed development, while that of tZ increased. We further analyzed the function of JcCYP735A using the CRISPR-Cas9 system, and found that the concentrations of tZ and tZ-riboside decreased significantly in the Jccyp735a mutants, which showed severely retarded growth. These findings will be helpful for further studies of the functions of cytokinin metabolic genes and understanding the roles of cytokinins in Jatropha growth and development.
Highlights
Jatropha curcas is a multipurpose tree that belongs to the Euphorbiaceae family
Orthologues of CKX1, 5, 6, and 7 formed a clade, while those of CKX2, 3, and 4 formed a separate clade (Fig. 5). These results showed that JcIPTs, JcCYP735A, and JcCKXs were most closely related to genes from R. communis, which belongs to the Euphorbiaceae family
Our study showed that JcCYP735A was highly expressed and that the tZ concentration increased with seed development in Jatropha (Table 1)
Summary
Jatropha curcas is a multipurpose tree that belongs to the Euphorbiaceae family. It can endure drought and adapt to barren land in tropical and subtropical regions. Jatropha is considered a promising biofuel plant due to the high oil content in its seeds (Akashi, 2012; Francis, Edinger & Becker, 2005; Makkar & Becker, 2009). The seed yield is very low, potentially because of the relatively low number of total flowers and/or the ratio of female to male flowers in Jatropha (Kumar & Sharma, 2008; Kumar Tiwari, Kumar & Raheman, 2007; Rao et al, 2008). Several studies have reported that exogenous cytokinin (CK) treatment can significantly increase the total number of flowers per inflorescence, the female-to-male flower ratio, and the seed yield (Froschle, Horn & Spring, 2017; Pan & Xu, 2011). The major derivatives are generally iP- and tZ-type CKs (Sakakibara, 2006)
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