Nitrate transporters (NRTs) are regulators of nitrate assimilation and transport.  The genome sequences of TaNRT2L12-A, -B and -D were cloned from wheat (Triticum aestivum L.), and polymorphisms were analyzed by sequencing.  TaNRT2L12-D in a germplasm population was highly conserved.  However, 38 single nucleotide polymorphisms (SNPs) in TaNRT2L12-A coding region and 11 SNPs in TaNRT2L12-B coding region were detected.  Two derived cleaved amplified polymorphic sequences (dCAPS) markers A-CSNP1 and A-CSNP2 were developed for TaNRT2L12-A based on SNP-351 and SNP-729, and three haplotypes were identified in the germplasm population.  B-CSNP1 and B-CSNP2 were developed for TaNRT2L12-B based on SNP-237 and SNP-1 227, and three haplotypes were detected in the germplasm population.  Association analyses between the markers and agronomic traits in 30 environments and phenotypic comparisons revealed that A-CSNP2-A is a superior allele of shorter plant height (PH), length of penultimate internode (LPI) and peduncle length (PL), B-CSNP2-G is a superior allele of higher grain number per spike (GNS).  Hap-6B-1 containing both superior alleles B-CSNP1-C and B-CSNP2-A is a superior haplotype of 1 000-grain weight (TGW).  Expression analysis showed that TaNRT2L12-B is mainly expressed in the root base and regulated by nitrate.  Therefore, TaNRT2L12 may be involved in nitrate transport and signaling to regulate TGW in wheat.  The superior alleles and dCAPS markers of TaNRT2L12-A/B are beneficial to genetic improvement and germplasm enhancement with molecular markers-assisted selection. 

 

Mealybugs, such as Phenacoccus solenopsis, are highly sexually dimorphic.  Winged adult males present such remarkable morphological differences from females that, to the untrained eye, conspecific adults of both sexes of P. solenopsis may be considered as two different insect species.  A method to investigate sex-dimorphic mechanisms is by evaluating gene expression using RT-qPCR.  However, the accuracy and consistency of this technique depend on the reference gene(s) selected.  In this study, we analyzed the expression of 10 candidate reference genes in male and female P. solenopsis at different development stages, using common algorithms including the ?Ct method, NormFinder, geNorm, BestKeeper, and a web-based analysis tool, RefFinder.  The results showed that EF1-β, RP-L32 and RP-18S were selected as the most stable genes by both the ?Ct method and NormFinder; TUB-α was the most stable gene identified by BestKeeper; and RP-L40 and RP-L32 were the most stable genes ranked by geNorm.  RefFinder, a comprehensive analysis software, ranked the ten genes and determined EF1-β and RP-L32 as the most suitable reference genes for the various developmental stages in male and female P. solenopsis.  Furthermore, the two most suitable reference genes were validated by examining expression of the juvenile hormone acid O-methytransferase (JHAMT) gene.  Results of the validation portion of the study showed that JHAMT expression was sex-biased towards males and exhibited a dynamic and classic expression pattern among the P. solenopsis developmental stages.  The results can help further our knowledge on the molecular mechanisms underlying sexual dimorphic development in P. solenopsis.