Spatial multi-omics at subcellular resolution via high-throughput in situ pairwise sequencing.

Technology for spatial multi-omics aids the discovery of new insights into cellular functions and disease mechanisms. Here we report the development and applicability of multi-omics in situ pairwise sequencing (MiP-seq), a method for the simultaneous detection of DNAs, RNAs, proteins and biomolecules at subcellular resolution. Compared with other in situ sequencing methods, MiP-seq enhances decoding capacity and reduces sequencing and imaging costs while maintaining the efficacy of detection of gene mutations, allele-specific expression and RNA modifications. MiP-seq can be integrated with in vivo calcium imaging and Raman imaging, which enabled us to generate a spatial multi-omics atlas of mouse brain tissues and to correlate gene expression with neuronal activity and cellular biochemical fingerprints. We also report a sequential dilution strategy for resolving optically crowded signals during in situ sequencing. High-throughput in situ pairwise sequencing may facilitate the multidimensional analysis of molecular and functional maps of tissues.

De novo genome assembly of a foxtail millet cultivar Huagu11 uncovered the genetic difference to the cultivar Yugu1, and the genetic mechanism of imazethapyr tolerance.

BACKGROUND: Setaria italica is the second-most widely planted species of millets in the world and an important model grain crop for the research of C4 photosynthesis and abiotic stress tolerance. Through three genomes assembly and annotation efforts, all genomes were based on next generation sequencing technology, which limited the genome continuity. RESULTS: Here we report a high-quality whole-genome of new cultivar Huagu11, using single-molecule real-time sequencing and High-throughput chromosome conformation capture (Hi-C) mapping technologies. The total assembly size of the Huagu11 genome was 408.37 Mb with a scaffold N50 size of 45.89 Mb. Compared with the other three reported millet genomes based on the next generation sequencing technology, the Huagu11 genome had the highest genomic continuity. Intraspecies comparison showed about 94.97 and 94.66% of the Yugu1 and Huagu11 genomes, respectively, were able to be aligned as one-to-one blocks with four chromosome inversion. The Huagu11 genome contained approximately 19.43 Mb Presence/absence Variation (PAV) with 627 protein-coding transcripts, while Yugu1 genomes had 20.53 Mb PAV sequences encoding 737 proteins. Overall, 969,596 Single-nucleotide polymorphism (SNPs) and 156,282 insertion-deletion (InDels) were identified between these two genomes. The genome comparison between Huagu11 and Yugu1 should reflect the genetic identity and variation between the cultivars of foxtail millet to a certain extent. The Ser-626-Aln substitution in acetohydroxy acid synthase (AHAS) was found to be relative to the imazethapyr tolerance in Huagu11. CONCLUSIONS: A new improved high-quality reference genome sequence of Setaria italica was assembled, and intraspecies genome comparison determined the genetic identity and variation between the cultivars of foxtail millet. Based on the genome sequence, it was inferred that the Ser-626-Aln substitution in AHAS was responsible for the imazethapyr tolerance in Huagu11. The new improved reference genome of Setaria italica will promote the genic and genomic studies of this species and be beneficial for cultivar improvement.

Synthesis, cytotoxicity, and in vivo antitumor activity study of parthenolide semicarbazones and thiosemicarbazones.

Parthenolide is an important sesquiterpene lactone with potent anticancer activities. In order to further improve its biological activity, a series of parthenolide semicarbazone or thiosemicarbazone derivatives was synthesized and evaluated for their anticancer activity. Derivatives were tested in vitro against 5 human tumor cell lines, and many of these showed higher cytotoxicity than parthenolide. Five compounds were further studied for their antitumor activity in mice. The in vivo result indicated that compound 4d showed both promising antitumor activity against mice colon tumor and small side effects on immune systems. The cell apoptosis and cell cycle distribution of compound 4d were also studied. Molecular docking studies revealed multiple interactions between 4d and NF-κB. Our findings demonstrate the potential of semicarbazones as a promising type of compounds with anticancer activity.

Efficient separation of vitamins mixture in aqueous solution using a stable zirconium-based metal-organic framework.

High-efficiency separation of niacin (NIA) and nicotinamide (NIC) still faces large challenge up to date. In this work, a stable zirconium-based metal-organic framework (DUT-67) was used to adsorb and separate NIA and NIC in aqueous solutions. The adsorption capacities for NIA and NIC at the concentration ratio of 1:1 were 110.2 mg g-1 and 11.2 mg g-1, respectively. Further study indicates that low ratio of NIA/ NIC is in favor of the separation. High temperature can restrain the adsorptions of NIA and NIC but promote the separation. Besides, DUT-67 can be well regenerated via a simple method. Mechanism analysis indicates that electrostatic interaction plays a critical role in the separation of these vitamins.

Synthetic Optimization of Ellipticine and Antitumor Activity of Novel Hexacyclic Derivatives of Ellipticine.

BACKGROUND: For decades, a great deal of research work has been done to synthesize ellipticine and its derivatives because of their potential antitumor properties and anti-HIV activities. However, the resonance structures in different media, a low level of solubility at physiological pH and systemic toxicity have prevented the use of ellipticine as a therapeutic agent. Besides, the low yield and complex steps of ellipticine synthesis limit its application. METHODS: A high-yield synthetic procedure of ellipticine has been optimized, and the total yield was up to 50% without silica gel column chromatography. Novel hexacyclic ellipticine derivatives were synthesized by coupling ellipticine with o-aminobenzoic acid. Their cytotoxicities against HCT116, MGC803, HT29 and MCF-7 tumor cells were evaluated. RESULTS: The synthesis process of ellipticine was optimized, and the total yield of the synthetic route was increased to 50% through several operation steps optimization. Fourteen ellipticine hexacyclic derivatives were synthesized. The synthetic compounds were screened for anti-tumor activity in vivo and in vitro, and some of the derivatives had good anti-tumor activity. CONCLUSION: Compared with ellipticine, the compound 1l showed higher antitumor activity and better tolerance to tumor models. The compound 1l treatment increased the percentage of late apoptotic cells from 3.1% (DMSO) to 21.6% (20.0 µM) in NCI-H460 cells. It also was observed the effect of 1l on G2 phase arrest was similar as that of ellipticine. The mechanism of action indicated compound 1l could be a topoisomerase IIα poison. These studies provided the basis for the pharmacodynamics and toxicology of ellipticine, and further clarifies the structureactivity relationship of antitumor activity of ellipticine.

Corticosterone impairs gap junctions in the prefrontal cortical and hippocampal astrocytes via different mechanisms.

Increasing evidence has implicated astrocyte pathology in the etiopathology of major depressive disorder (MDD). In particular, dysfunction of gap junctions in astrocytes is a potential target for MDD treatment. However, the mechanism underlying stress-induced dysfunction of gap junctions is still unknown. We therefore studied the mechanism of stress-induced dysfunction of gap junctions in prefrontal cortical and hippocampal astrocytes. Corticosterone (CORT) was used to induce stress conditions; CORT damaged the function of gap junctions, which resulted from less distribution of connexin43 (Cx43) on membranes and the enhanced phosphorylation of Cx43 at S368. Moreover, CORT downregulated the biosynthesis of Cx43 but increased the degradation of Cx43. Interestingly, both autophagy and the proteasome system were involved in the degradation of Cx43 in prefrontal cortical astrocytes, but only the proteasome system was involved in the degradation of Cx43 in hippocampal astrocytes. CORT significantly induced the formation of annular gap junction vesicles in prefrontal cortical astrocytes; however, Cx43 mainly presented as small dots in the hippocampal astrocytes. Furthermore, CORT increased N-Cadherin expression and the interactions of Cx43 with ZO-1/drebrin in prefrontal cortical astrocytes, but these interactions were oppositely modulated in hippocampal astrocytes. In conclusion, this study clarified the alternations of the Cx43 life cycle in the prefrontal cortical and hippocampal astrocytes exposed to CORT, which may contribute to our understanding of the mechanisms underlying stress-induced dysfunction of gap junctions.

Taxonomic structure and functional association of foxtail millet root microbiome.

The root microbes play pivotal roles in plant productivity, nutrient uptakes, and disease resistance. The root microbial community structure has been extensively investigated by 16S/18S/ITS amplicons and metagenomic sequencing in crops and model plants. However, the functional associations between root microbes and host plant growth are poorly understood. This work investigates the root bacterial community of foxtail millet (Setaria italica) and its potential effects on host plant productivity. We determined the bacterial composition of 2882 samples from foxtail millet rhizoplane, rhizosphere and corresponding bulk soils from 2 well-separated geographic locations by 16S rRNA gene amplicon sequencing. We identified 16 109 operational taxonomic units (OTUs), and defined 187 OTUs as shared rhizoplane core OTUs. The ß-diversity analysis revealed that microhabitat was the major factor shaping foxtail millet root bacterial community, followed by geographic locations. Large-scale association analysis identified the potential beneficial bacteria correlated with plant high productivity. Besides, the functional prediction revealed specific pathways enriched in foxtail millet rhizoplane bacterial community. We systematically described the root bacterial community structure of foxtail millet and found its core rhizoplane bacterial members. Our results demonstrated that host plants enrich specific bacteria and functions in the rhizoplane. The potentially beneficial bacteria may serve as a valuable knowledge foundation for bio-fertilizer development in agriculture.

[Anti-tumor target prediction and activity verification of Ganoderma lucidum triterpenoids].

It has reported that Ganoderma lucidum triterpenoids had anti-tumor activity. However, the anti-tumor target is still unclear. The present study was designed to investigate the anti-tumor activity of G. lucidum triterpenoids on different tumor cells, and predict their potential targets by virtual screening. In this experiment, molecular docking was used to simulate the interactions of 26 triterpenoids isolated from G. lucidum and 11 target proteins by LibDock module of Discovery Studio2016 software, then the anti-tumor targets of triterpenoids were predicted. In addition, the in vitro anti-tumor effects of triterpenoids were evaluated by MTT assay by determining the inhibition of proliferation in 5 tumor cell lines. The docking results showed that the poses were greater than five, and Libdock Scores higher than 100, which can be used to determine whether compounds were activity. Eight triterpenoids might have anti-tumor activity as a result of good docking, five of which had multiple targets. MTT experiments demonstrated that the ganoderic acid Y had a certain inhibitory activity on lung cancer cell H460, with IC50 of 22.4 µmol•L ⁻¹, followed by 7-oxo-ganoderic acid Z2, with IC50 of 43.1 µmol•L ⁻¹. However, the other triterpenoids had no anti-tumor activity in the detected tumor cell lines. Taking together, molecular docking approach established here can be used for preliminary screening of anti-tumor activity of G.lucidum ingredients. Through this screening method, combined with the MTT assay, we can conclude that ganoderic acid Y had antitumor activity, especially anti-lung cancer, and 7-oxo-ganoderic acid Z2 as well as ganoderon B, to a certain extent, had anti-tumor activity. These findings can provide basis for the development of anti-tumor drugs. However, the anti-tumor mechanisms need to be further studied.

Ginsenoside Rg1 alleviates corticosterone-induced dysfunction of gap junctions in astrocytes.

ETHNOPHARMACOLOGICAL RELEVANCE: Ginsenoside Rg1 (Rg1), one of the major bioactive ingredients of Panax ginseng C. A. Mey, has neuroprotective effects in animal models of depression, but the mechanism underlying these effects is still largely unknown AIM OF THE STUDY: Gap junction intercellular communication (GJIC) dysfunction is a potentially novel pathogenic mechanism for depression. Thus, we investigated that whether antidepressant-like effects of Rg1 were related to GJIC. MATERIALS AND METHODS: Primary rat prefrontal cortical and hippocampal astrocytes cultures were treated with 50µM CORT for 24h to induce gap junction damage. Rg1 (0.1, 1, or 10µM) or fluoxetine (1µM) was added 1h prior to CORT treatment. A scrape loading and dye transfer assay was performed to identify the functional capacity of gap junctions. Western blot was used to detect the expression and phosphorylation of connexin43 (Cx43), the major component of gap junctions. RESULTS: Treatment of primary astrocytes with CORT for 24h inhibited GJIC, decreased total Cx43 expression, and increased the phosphorylation of Cx43 at serine368 in a dose-dependent manner. Pre-treatment with 1µM and 10µM Rg1 significantly improved GJIC in CORT-treated astrocytes from the prefrontal cortex and hippocampus, respectively, and this was accompanied by upregulation of Cx43 expression and downregulation of Cx43 phosphorylation. CONCLUSION: These findings provide the first evidence indicating that Rg1 can alleviate CORT-induced gap junction dysfunction, which may have clinical significance in the treatment of depression.

Identification of QTLs for 14 Agronomically Important Traits in Setaria italica Based on SNPs Generated from High-Throughput Sequencing.

Foxtail millet (Setaria italica) is an important crop possessing C4 photosynthesis capability. The S. italica genome was de novo sequenced in 2012, but the sequence lacked high-density genetic maps with agronomic and yield trait linkages. In the present study, we resequenced a foxtail millet population of 439 recombinant inbred lines (RILs) and developed high-resolution bin map and high-density SNP markers, which could provide an effective approach for gene identification. A total of 59 QTL for 14 agronomic traits in plants grown under long- and short-day photoperiods were identified. The phenotypic variation explained ranged from 4.9 to 43.94%. In addition, we suggested that there may be segregation distortion on chromosome 6 that is significantly distorted toward Zhang gu. The newly identified QTL will provide a platform for sequence-based research on the S. italica genome, and for molecular marker-assisted breeding.

Updated foxtail millet genome assembly and gene mapping of nine key agronomic traits by resequencing a RIL population.

Foxtail millet (Setaria italica) provides food and fodder in semi-arid regions and infertile land. Resequencing of 184 foxtail millet recombinant inbred lines (RILs) was carried out to aid essential research on foxtail millet improvement. A total 483 414 single nucleotide polymorphisms were determined. Bin maps were constructed based on the RILs' recombination data. Based on the high-density bin map, we updated Zhanggu reference with 416 Mb after adding 16 Mb unanchored scaffolds and Yugu reference with some assembly error correction and 3158 gaps filled. Quantitative trait loci (QTL) mapping of nine agronomic traits was done based on this RIL population, five of which were controlled by a single gene. Meanwhile, two QTLs were found for plant height, and a candidate gene showed 89% identity to the known rice gibberellin-synthesis gene sd1. Three QTLs were found for the trait of heading date. The whole genome resequencing and QTL mapping provided important tools for foxtail millet research and breeding. Resequencing of the RILs could also provide an effective way for high-quality genome assembly and gene identification.

Ginsenoside Rg1-induced antidepressant effects involve the protection of astrocyte gap junctions within the prefrontal cortex.

Ginsenoside Rg1 (Rg1) exhibits antidepressant-like activity by increasing neurogenesis and dendritic spine density without discernible side effects. However, the molecular mechanisms underlying Rg1 antidepressant activity remain poorly understood. As the dysfunction of gap junctions between astrocytes in the prefrontal cortex (PFC) is implicated in major depression disorder, the aim of this study was to investigate the effects of Rg1 on astrocyte gap junctions in the PFC. Rats exposed to chronic unpredictable stress (CUS) were administered Rg1 (5, 10, and 20mg/kg) for 28days and analyzed for depressive symptoms using the sucrose preference and forced swimming tests. Functional and morphological changes of gap junction channels in the PFC were evaluated using dye transfer and electron microscopy, respectively. The expression of connexin 43 (Cx43) was analyzed by western blotting. Rg1 markedly alleviated depression-like behavior in rats. Long-term Rg1 treatment of CUS-exposed rats also significantly prevented the decrease in dye diffusion and improved the ultrastructure of astrocyte gap junctions in the PFC, indicating beneficial effects on the functional activity of gap junction channels in the brain. In addition, Rg1 upregulated Cx43 expression in the PFC reduced by CUS exposure, which significantly correlated with its antidepressant-like effects. The results demonstrate that Rg1-induced antidepressant effects are might be mediated, in part, by protecting astrocyte gap junctions within the prefrontal cortex.

[The basic strategies and research advances in the studies on glycosyltransferases involved in ginsenoside biosynthesis].

Traditional herbal medicines, Panax ginseng, Panax quinquefolium and Panax notoginseng, attract our attention for their extensive and powerful pharmacological activities. Ginsenosides are the main active constituents of these medicinal herbs. The related glycosyltransferases involved in ginsenoside biosynthesis are the key enzymes which catalyze the last important step. Modification of ginsenoside aglycones by glycosyltransferases produces the complexity and diversity of ginsenosides, which have more extensive pharmacological activity. At present, ginsenoside aglycones and compound K have been obtained by synthetic biology. As the last step of ginsenoside biosynthesis, glycosylation of ginsenoside aglycones has been studied intensively in recent years. This review summarizes the basic strategies and research advances in studies on glycosyltransferases involved in ginsenoside biosynthesis, which is expected to lay the theoretical foundation for the in-depth research of biosynthetic pathway of ginsenosides and their production by synthetic biology.

[Downregulation of lanosterol synthase gene expression by antisense RNA technology in Saccharomyces cerevisiae].

The cyclization of 2,3-oxidosqualene is the key branch point of ergosterol and triterpenoid biosynthesis. Downregulation of 2,3-oxidosqualene metabolic flux to ergosterol in Saccharomyces cerevisiae may redirect the metabolic flux toward the triterpenoid synthetic pathway. In our study, primers were designed according to erg7 gene sequence of S. cerevisiae. Three fragments including 5' long fragment, 5' short fragment and erg7 coding region fragment were amplified by PCR. 5' long fragment consists of the promoter and a part of erg7 coding region sequence. 5' short fragment consists of a part of promoter and a part of erg7 coding region sequence. These fragments were inserted reversely into pESC-URA to construct antisense expression plasmids. The recombinant plasmids were transformed into S. cerevisiae INVSc1 and recombinant strains were screened on the nutritional deficient medium SD-URA. The erg7 expression level of recombinant strains, which harbored antisense expression plasmid of erg7 coding region, was similar to that of INVScl by semi-quantitative PCR detection. But erg7 expression level of recombinant strains, which harbored 5' long antisense fragment and 5' short antisense fragment, was significantly lower than that of the control. The results of TLC and HPLC showed that the ergosterol content of recombinant strains, which harbored 5' long antisense fragment, decreased obviously. The ergosterol contents of the others were almost equal to that of INVSc1. Lanosterol synthase gene expression was downregulated by antisense RNA technology in S. cerevisiae, which lays a foundation for reconstructing triterpenoid metabolic pathway in S. cerevisiae by synthetic biology technology.

Prokaryotic ubiquitin-like ThiS fusion enhances the heterologous protein overexpression and aggregation in Escherichia coli.

Fusion tags are commonly employed to enhance target protein expression, improve their folding and solubility, and reduce protein degradation in expression of recombinant proteins. Ubiquitin (Ub) and SUMO are highly conserved small proteins in eukaryotes, and frequently used as fusion tags in prokaryotic expression. ThiS, a smaller sulfur-carrier protein involved in thiamin synthesis, is conserved among most prokaryotic species. The structural similarity between ThiS and Ub provoked us into expecting that the former could be used as a fusion tag. Hence, ThiS was fused to insulin A and B chains, murine Ribonuclease Inhibitor (mRI) and EGFP, respectively. When induced in Escherichia coli, ThiS-fused insulin A and B chains were overexpressed in inclusion bodies, and to higher levels in comparison to the same proteins fused with Ub. On the contrast, ThiS fusion of mRI, an unstable protein, resulted in enhanced degradation that was not alleviated in protease-deficient strains. While the degradation of Ub- and SUMO-fused mRI was less and seemed protease-dependent. Enhanced degradation of mRI did not occur for the fusions with half-molecules of ThiS. When ThiS-tag was fused to the C-terminus of EGFP, higher expression, predominantly in inclusion bodies, was observed again. It was further found that ThiS fusion of EGFP significantly retarded its refolding process. These results indicated that prokaryotic ThiS is able to promote the expression of target proteins in E. coli, but enhanced degradation may occur in case of unstable targets. Unlike eukaryotic Ub-based tags usually increase the solubility and folding of proteins, ThiS fusion enhances the expression by augmenting the formation of inclusion bodies, probably through retardation of the folding of target proteins.

(+)-epi-Clausenamide, but not (-)-epi-clausenamide, showed more potential than (-)-clausenamide on facilitating synaptic transmission in CA1 region of hippocampal synapses.

Effect of (+)-epi-clausenamide and (-)-epi-clausenamide on synaptic transmission in CA1 region of hippocampal slice was compared in this study. (+)-epi-Clausenamide showed more potency than (-)-clausenamide on either induction or maintenance of long term potentiation (LTP). Effect of (+)-epi-clausenamide on potentiating basic synaptic transmission was also superior to (-)-clausenamide. However, (-)-epi-clausenamide showed only slight effects on synaptic transmission, suggesting that the effect of (+)-epi-clausenamide and (-)-epi-clausenamide on synaptic transmission depended on their chirality. Calcium influx was not involved in (+)-epi-clausenamide facilitating synaptic transmission in this study. (+)-epi-Clausenamide might promote glutamate release through the activation of Synapsin I(Ser9) to activate the downstream effectors which play a key role in synaptic plasticity. Elucidating the mechanism of each optical isomer of clausenamide by electrophysiological methods provided basis for therapeutic strategies for neurological disorders.

Overexpression of alpha-synuclein down-regulates BDNF expression.

Parkinson's disease (PD) is a chronic progressive neurodegenerative movement disorder characterized by the selective loss of nigrostriatal dopaminergic neurons. However, the molecular pathways leading to the dopaminergic neuron degeneration have remained obscure until recently. Reports demonstrated that reduction of brain-derived neurotrophic factor (BDNF) was involved in the etiology and pathogenesis of PD, but its mechanism has not been elucidated. alpha-Synuclein has a causal role in Parkinson's disease, and could interfere with transcriptional regulation of dopamine neurons. In this study, alpha-synuclein overexpression was found to decrease the expression of BDNF, and also to suppress the transactivation of nuclear factors of activated T-cells (NFAT) and cAMP response element binding protein (CREB), both of which regulate BDNF expression. Furthermore, overexpressed alpha-synuclein could associate with protein kinase C (PKC) and impair its activity. Meanwhile glycogen synthase kinase-3beta (GSK3beta) was activated and extracellular signal-regulated protein kinase (ERK) activity was inhibited by overexpression of alpha-synuclein; both of them were downstream kinases of PKC. Therefore, the impaired PKC signal pathway caused by alpha-synuclein overexpression might account at least partially for the down-regulation of BDNF.

[Elucidating the structure of two cyclotides of Viola tianshanica maxim by MALDI TOF/TOF MS analysis].

The cyclotides are a family of cyclic "mini" proteins that occur in Violaceae, Rubiaceae and Cucurbitaceae plant families and contain a head-to-tail cyclic backbone and a cystine knot arranged by three disulfide bonds. To study the natural cyclotides of V tianshanica, dried herb was extracted with 50% ethanol, and the concentrated aqueous extract was subjected to a solvent-solvent partitioning between water and hexane, ethyl acetate and n-butanol, separately. The n-butanol extract containing cyclotides was subjected to column chromatography over Sephadex LH-20, eluted with 30% methanol. The subfractions were directly reduced by DTT and analyzed by reverse-phase HPLC. The peaks with different retention times were shown on the profile of RP-HPLC and collected. The cyclotides were speculated based on masses range from 3 000 to 3 500 Da. The purified cyclotides were reduced with DTT, alkylated with iodoacetamide, and then were cleaved with endoproteinase Glu-C, endoproteinase Lys-C and Trypsin, separately. The digested peptides were purified on RP-HPLC and analyzed on MALDI TOF/TOF analyzer. A new cyclotide, cycloviolacin T1 and a reported cyclotide varv E were systemically determined using MALDI TOF/TOF system. So the method for the isolation and characterization of cyclotides was quickly built up in succession.

[Characteristics of references between GSCC and CST for Semi-Luvisols].

In this paper, the references between Genetic Soil Classification of China (GSCC) and the Chinese Soil Taxonomy (CST) for GSCC-Semi-Luvisols were conducted, and their quantitative and spatial distribution characteristics within CST were studied, based on the 1 : 1 M Soil Database of China, which consists of 1 : 1 M digital soil map, soils profiles attribution database and soil reference system of China. Being a reference system for converting soil names in GSCC into those in CST, ST and WRB, respectively, Chinese Soil Reference System was a computerized retrieving system jointly developed by the experienced scientists of pedology and computer science. The comparison fields and laboratory investigation data of their soil profiles with diagnostic horizons and characteristics related in the target soil classification systems, and 2,540 typical soil species names corresponding in CST, ST and WRB systems were determined, respectively, which were selected from Soil Attributes Database because of their complete sets of attribute data. Finally, the system and reference database were established. "GIS linkage based soil type" method linked the records in the Soil Reference Database to the Soil Spatial Database. In this method, all records of soil profiles in Soil Reference Database as well as their soil reference name in other classification systems were allocated one by one onto corresponding soil type polygons in Soil Spatial Database on the GIS platform, according to the principles of soil type identity and similarity, parent material identity and likeness, and the location of soil profiles relative to linked target polygons. Area statistcs of all soils were conducted based on the polygons. The results showed that GSCC-Semi-Luvisols was a type of GSCC soil with a total area of 427,843.1 km2,which could be sorted to 4 CST Orders, i. e., Luvisols (51.3%), Cambosols (35.2%), Isohumosols (10.7%) and Anthrosols (2.8%), and further into CST 22 Groups and 38 Subgroups. All dark grey forest soil, superficial gleyed black soil, and leached dry red soil of GSCC subgroups could be sorted to Calcaric Hapli-Gelic Cambosols, Pachic Argi-Udic Cambosols and Typic Ferri-Ustic Luvisols of CST subgroups, respectively, and all grey cinnamon-like soil, calcareous grey cinnamonic soil and dry cinnamon soil could be sorted to Typic Ustic Cambosols. Making the reference was so complicated that there was no one to one reference relationship among other soils. The analysis of the area ratios and standard deviations of a certain GSCC soil classified by CST showed that the lower the unit for reference, the easier the reference would be. The results of this study were of high reference value to proper reference GSCC and CST, and to the application and development of CST.