Thrips hawaiiensis (Morgan, 1913) is an important insect pest of fruits and vegetables. At present, it is primarily distributed in tropical and subtropical area. For a better understanding of the genetic makeup and migration patterns of T. hawaiiensis throughout the world, we isolated 11 novel polymorphic microsatellite loci from an enriched genomic library based on a biotin/streptavidin capture protocol. Genetic parameters were estimated on 80 individual thrips from two natural populations. The results showed that all 11 loci were highly polymorphic; the number of alleles ranged from six to 37, and nine loci demonstrated polymorphic information content (PIC) > 0.5. The observed (H
 O) and expected (H
 E) heterozygosities ranged from 0.350 to 0.925 and 0.307 to 0.952, respectively. Furthermore, only four locus/population combinations significantly deviated from Hardy–Weinberg equilibrium (HWE). These microsatellite markers have potential utility in population structure and gene flow studies of this species.

Polymorphic microsatellite markers in Thrips hawaiiensis (Thysanoptera: Thripidae)

Cryptolestes ferrugineus (Stephens, 1831) is an important insect pest of stored products. Due to its broad host range, short life cycle, and high reproductive capacity, this species has rapidly colonized temperate and tropical regions around the world. In this study, we isolated 18 novel polymorphic microsatellite loci from an enriched genomic library based on a biotin/streptavidin capture protocol. These loci will be useful tool to better understand the genetic structure and migration patterns of C. ferrugineus throughout the world. The genetic parameters were estimated based on 80 individual C. ferrugineus from two natural populations. The results revealed that 18 loci were different polymorphic levels. The numbers of alleles ranged from 3 to 12, and eleven loci demonstrated polymorphic information contents greater than 0.5. The observed (H O) and expected (H E) heterozygosities ranged from 0.051 to 0.883 and 0.173 to 0.815, respectively. Five locus/population combinations significantly deviated from Hardy-Weinberg equilibrium. We also demonstrated the potential utility of the C. ferrugineus microsatellites as population and species markers for four additional Cryptolestes species.

Microsatellite markers for Cryptolestes ferrugineus (Coleoptera: Laemophloeidae) and other Cryptolestes species.

Ovule abortion is a common phenomenon in plants that has an impact on seed production. Previous studies of ovule and female gametophyte (FG) development have mainly focused on angiosperms, especially in Arabidopsis thaliana. However, because it is difficult to acquire information about ovule development in gymnosperms, this remains unclear. Here, we investigated the transcriptomic data of natural ovule abortion mutants (female sterile line, STE) and the wild type (female fertile line, FER) of Pinus tabuliformis Carr. to evaluate the mechanism of ovule abortion during the process of free nuclear mitosis (FNM). Using single-molecule real-time (SMRT) sequencing and next-generation sequencing (NGS), 18 cDNA libraries via Illumina and two normalized libraries via PacBio, with a total of almost 400,000 reads, were obtained. Our analysis showed that the numbers of isoforms and alternative splicing (AS) patterns were significantly variable between FER and STE. The functional annotation results demonstrate that genes involved in the auxin response, energy metabolism, signal transduction, cell division, and stress response were differentially expressed in different lines. In particular, AUX/IAA, ARF2, SUS, and CYCB had significantly lower expression in STE, showing that auxin might be insufficient in STE, thus hindering nuclear division and influencing metabolism. Apoptosis in STE might also have affected the expression levels of these genes. To confirm the transcriptomic analysis results, nine pairs were confirmed by quantitative real-time PCR. Taken together, these results provide new insights into ovule abortion in gymnosperms and further reveal the regulatory mechanisms of ovule development.

https://www.mdpi.com/1422-0067/22/6/3138/pdf

Combined Transcriptome Analysis Reveals the Ovule Abortion Regulatory Mechanisms in the Female Sterile Line of Pinus tabuliformis Carr.

The invention provides a DNA extracting method for bitter gourd hybrid purity molecular identification. The method comprises the following steps: on the basis of extracting genome DNA through an improved CTAB method, respectively feeding a large number of bitter gourd tender leaves from different plants required for detecting in 1.5ml centrifugal tubes, placing the centrifugal tubes in a refrigerator at -80 DEG C for more than 3 hours, taking out, grinding about 5s by using a cleaned grinding pestle, quickly adding 2*CTAB extracting buffer solution, and then suspending through chloroform/isoamylol, washing and precipitating through isopropanol and 70% ethanol to obtain a large number of to-be-detected sample bitter gourd genome DNA of different plants. The problem that the step of extracting the bitter gourd genome DNA by using the traditional CTAB method is excessively fussy is solved; and meanwhile, the different to-be-detected bitter gourd genome DNA obtained through extracting is basically the same in concentration, and can be completely suitable for molecular identification of RAPD, SRAP and ISSR of bitter gourd hybrid purity in quality and completeness.

DNA extracting method for bitter gourd hybrid purity molecular identification

To monitor and control the Japanese orange fly Bactrocera tsuneonis (Miyake) (Diptera: Tephritidae), it is important to determine their dispersal ability. Therefore, to determine the genetic structure of this species, we developed microsatellite markers at 17 loci for B. tsuneonis. A total of 80 adult females and males randomly collected from abandoned orchards in the Oita and Yamaguchi Prefectures, in the western part of Japan, were used for polymorphism analysis using the microsatellite markers. The average number of alleles was 2.65 for the 17 loci, in which two loci did not show polymorphism. The expected heterozygosity ranged from 0 to 0.663, although the observed heterozygosity was relatively low between 0 and 0.105. The null allele frequencies varied among populations. Moreover, no polymorphism was found in the three larval populations collected in the Oita Prefecture. These results suggested a high degree of inbreeding for this species. Significant genetic differentiation between females and males of the same population was not observed, although it was observed between the Oita and Yamaguchi populations. A bottleneck effect was detected in Yamaguchi but not in Oita, and this is congruent with the history of an expanding distribution in this species. These markers could be used for population discrimination and to expand the geographic history of this species, potentially contributing to the pest management of B. tsuneonis. 

/pdf/development-of-microsatellite-markers-for-the-japanese-eeemotsm.pdf

Development of microsatellite markers for the Japanese orange fly, Bactrocera tsuneonis (Diptera: Tephritidae)

Bactrocera correcta (Bezzi) is one of the most destructive insect pests of fruits and vegetables in tropical and subtropical regions. At present, this fly is primarily distributed in Southeast Asia. Twelve microsatellite loci were isolated from an enriched genomic library based on a biotin/streptavidin capture protocol. The polymorphism of these loci was tested on 74 individual flies from two natural populations. Allele number ranged from 6 to 14 and 10 loci demonstrated a polymorphic information content (PIC) greater than 0.5. The pairwise F
 ST value between the two populations was 0.0048 (P > 0.05). These microsatellite loci have potential utility for studies of population genetic structure in this species.

Polymorphic microsatellite markers in the guava fruit fly, Bactrocera correcta (Diptera: Tephritidae)

1Institute of Plant Quarantine, Chinese Academy of Inspection and Quarantine, Beijing, 100029, People’s Republic of China 2Gangxi Entry-Exit Inspection and Quarantine Bureau, Gangxi 530028, People’s Republic of China 3Department of Entomology, College of Agronomy and Biotechnology, China Agricultural University, Beijing 100093, People’s Republic of China 4State Key Laboratory of Tree Genetics and Breeding, The Research Institute of Forestry, Chinese Academy of Forestry, Beijing 100091, People’s Republic of China

Polymorphic microsatellite markers in Taxus chinensis var. mairei (Taxaceae).

The invention establishes and optimizes an SSR-PCR reaction system of Taxus chinensis var. mairei. Through optimization tests on influence factors (primer, dNTP, Taq DNA polymerase, Mg and template DNA) of Taxus chinensis var. mairei SSR-PCR reaction system at multiple levels, the optimal levels of various reaction factors are screened to establish an optimal Taxus SSR-PCR 20 muL reaction system: 2 muM of primer, 40 nuM of dNTP, 1U of Taq DNA polymerase, 1.0 mM of Mg and 75 ng of template DNA. The establishment of the system can well meet the requirements of Taxus chinensis var. mairei genome DNA amplification, and lays foundation for SSR molecular marker and genetic polymorphism analysis of Taxus chinensis var. mairei.

Taxus chinensis var. mairei SSR-PCR reaction system and application thereof

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Polymorphic microsatellite markers in the guava fruit fly, Bactrocera correcta (Diptera: Tephritidae)

Polymorphic microsatellite markers in Taxus chinensis var. mairei (Taxaceae).

Taxus chinensis var. mairei SSR-PCR reaction system and application thereof