Genome editing is an important tool for gene functional studies as well as crop improvement. The recent development of the CRISPR/Cas9 system using single guide RNA molecules (sgRNAs) to direct precise double strand breaks in the genome has the potential to revolutionize agriculture. Unfortunately, not all sgRNAs are equally efficient and it is difficult to predict their efficiency by bioinformatics. In crops such as cotton (Gossypium hirsutum L.), with labor-intensive and lengthy transformation procedures, it is essential to minimize the risk of using an ineffective sgRNA that could result in the production of transgenic plants without the desired CRISPR-induced mutations. In this study, we have developed a fast and efficient method to validate the functionality of sgRNAs in cotton using a transient expression system. We have used this method to validate target sites for three different genes GhPDS, GhCLA1, and GhEF1 and analyzed the nature of the CRISPR/Cas9-induced mutations. In our experiments, the most frequent type of mutations observed in cotton cotyledons were deletions (∼64%). We prove that the CRISPR/Cas9 system can effectively produce mutations in homeologous cotton genes, an important requisite in this allotetraploid crop. We also show that multiple gene targeting can be achieved in cotton with the simultaneous expression of several sgRNAs and have generated mutations in GhPDS and GhEF1 at two target sites. Additionally, we have used the CRISPR/Cas9 system to produce targeted gene fragment deletions in the GhPDS locus. Finally, we obtained transgenic cotton plants containing CRISPR/Cas9-induced gene editing mutations in the GhCLA1 gene. The mutation efficiency was very high, with 80.6% of the transgenic lines containing mutations in the GhCLA1 target site resulting in an intense albino phenotype due to interference with chloroplast biogenesis.

BackgroundNLRPs (Nucleotide-binding oligomerization domain, Leucine rich Repeat and Pyrin domain containing Proteins) are members of NLR (Nod-like receptors) protein family. Recent researches have shown that NLRP genes play important roles in both mammalian innate immune system and reproductive system. Several of NLRP genes were shown to be specifically expressed in the oocyte in mammals. The aim of the present work was to study how these genes evolved and diverged after their duplication, as well as whether natural selection played a role during their evolution.ResultsBy using in silico methods, we have evaluated the evolution and functional divergence of NLRP genes, in particular of mouse reproduction-related Nlrp genes. We found that (1) major NLRP genes have been duplicated before the divergence of mammals, with certain lineage-specific duplications in primates (NLRP7 and 11) and in rodents (Nlrp1, 4 and 9 duplicates); (2) tandem duplication events gave rise to a mammalian reproduction-related NLRP cluster including NLRP2, 4, 5, 7, 8, 9, 11, 13 and 14 genes; (3) the function of mammalian oocyte-specific NLRP genes (NLRP4, 5, 9 and 14) might have diverged during gene evolution; (4) recent segmental duplications concerning Nlrp4 copies and vomeronasal 1 receptor encoding genes (V1r) have been undertaken in the mouse; and (5) duplicates of Nlrp4 and 9 in the mouse might have been subjected to adaptive evolution.ConclusionIn conclusion, this study brings us novel information on the evolution of mammalian reproduction-related NLRPs. On the one hand, NLRP genes duplicated and functionally diversified in mammalian reproductive systems (such as NLRP4, 5, 9 and 14). On the other hand, during evolution, different lineages adapted to develop their own NLRP genes, particularly in reproductive function (such as the specific expansion of Nlrp4 and Nlrp9 in the mouse).

This paper develops structural dynamic methods to project future carbon fluxes in forests. These methods account for land management changes on both the intensive and extensive margins, both of which are critical components of future carbon fluxes. When implemented, the model suggests that U.S. forests remain a carbon sink through most of the coming century, sequestering 128 Tg C y −1 . Constraining forestland to its current boundaries and constraining management to current levels reduce average sequestration by 25 to 28 Tg C y −1 . An increase in demand leads to increased management and greater sequestration in forests. The results are robust to climate change. (JEL Q23, Q54)

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Stress-associated proteins (SAPs) containing the A20/AN1 zinc-finger domain play important roles in response to both biotic and abiotic stresses in plants. Nevertheless, few studies have focused on the SAP gene family in cotton. To explore the distributions and expression patterns of these genes, we performed genome-wide identification and characterization of SAPs in tetraploid Gossypium hirsutum L. TM-1 (AD1). A total of 37 genes encoding SAPs were identified, 36 of which were duplicated in the A and D sub-genomes. The analysis of gene architectures and conserved protein motifs revealed that nearly all A20-AN1-type SAPs were intron-free, whereas AN1-AN1-type SAPs contained one intron. The cis-elements of the SAP promoters were studied, as were the expression levels of cotton SAP genes under different stresses based on RNA-seq data and validated by qRT-PCR. Most cotton SAP genes were induced by multiple stresses and phytohormones, particularly salt stress, indicating that SAP genes may play important roles in cotton's response to unfavorable environmental changes. Among these identified SAPs, the expression of GhSAP17A/D is suppressed in cotton response to Vertillium dahliae, and the GhSAP17A/D-silenced cotton exhibits more resistance to V. dahliae. This study provides insight into the evolution of SAP genes in upland cotton and may aid in efforts at further functional identification of A20/AN1-type proteins and cotton's response to different stresses.

This paper develops an economic analysis of climate change impacts in the global forest sector. It illustrates how potential future climate change impacts can be integrated into a dynamic forestry economics model using data from a global dynamic vegetation model, the MC2 model. The results suggest that climate change will cause forest outputs (such as timber) to increase by approximately 30% over the century. Aboveground forest carbon storage also is projected to increase, by approximately 26 Pg C by 2115, as a result of climate change, potentially providing an offset to emissions from other sectors. The effects of climate mitigation policies in the energy sector are then examined. When climate mitigation in the energy sector reduces warming, we project a smaller increase in forest outputs over the timeframe of the analysis, and we project a reduction in the sink capacity of forests of around 12 Pg C by 2115.

The avian egg, which contains the egg yolk, the egg white, and the eggshell, represents the mostly advanced amniotic egg in oviparous vertebrates. In mammals, this reproductive strategy of laying egg has gradually evolved toward placentation. In order to better understand the unique status of the avian egg in the evolution of the vertebrate reproduction, we investigated the evolution of some Gallus gallus egg-specific protein-coding genes. Based on our finding and other recent research, we have summarized here that gene formation (such as ovalbumin genes, ovocalyxin-36 and apovitellenin-1 encoding genes in the G. gallus), gene divergence between G. gallus and mammals (such as the ovocalyxin-32 gene with its ortholog, the mammalian RARRES1, and the ovocleidin-116 with its ortholog, the mammalian MEPE), and gene loss (egg-expressed genes lost during the evolution of the mammals, such as vitellogenin and RBP encoding genes) play significant roles in the evolution of egg-specific genes.

The Chinese agricultural sector has experienced a substantial increase in total output since dramatic reforms were introduced in 1978. This paper uses the index method to measure agricultural total factor productivity (TFP) for China’s crop and livestock industries, based on the gross output model from 1978 to 2016. We construct production accounts for the industries using input‐output relationships for the 26 main agricultural commodities and commodity groups, which account for over 90 per cent of the total agricultural inputs and outputs. The results show that China’s agricultural TFP grew at a rate of approximately 2.4 per cent a year before 2009, which is comparable to the main OECD countries and is double the world average. TFP growth accounts for approximately 40 per cent of output growth, suggesting that input growth was the main driver of output growth in the past. However, average productivity growth slowed down after 2009 though it has gradually recovered since 2012. The slowdown reflects the emerging challenges to existing farm production practices in Chinese agriculture, suggesting the need for further institutional reform.