High-level Expression and Purification of Human Zona Pellucida huZP3a^(22-176) and huZP3b^(177-348) Peptides in Escherichia coli

Objective To try making huZP3a^22-176 and huZP3b^177-348 polypeptides （representing an intact huZP^322-348 protein without its N-terminal signal peptide and C-terminal transmembrane domain ） express in E. coli at a higher level Methods The cDNAs encoding huZP3a and huZP3b were obtained with PCR method. The pBV221 plasmid was used to construct thermo-inducible recombinant expression vector. Purification of two target expression products employed an improved method of preparative gel polyacrylamide gel electrophoresis. Results Two polypeptides of recombinant huZP3a （rhuZP3a） and recombinant huZP3b （rhuZP3b） were all expressed respectively in an E. coli BL21（DE3）pLysS strain at a higher level, which were recognized by two specific polyclonal antisera in Western blotting test which recognize a linear B cell epitope present in rhuZP3a or rhuZP3b respectively. Using the shake-flask method, approximately 5 mg of rhuZP3a and rhuZP3b with more than 95% relative homogeneity were harvested from 1 L culture respectively. Conclusion The availability of two rhuZP3 polypeptides will help in detecting the immunogenicities of rhuZP3a and rhuZP3b through animal experiments and confirming the function domain of non-glycosylated huZP3 to induce acrosome reaction in vitro.

Optical scheme to demonstrate state-independent quantum contextuality

The contradiction between classical and quantum physics can be identified through quantum contextuality, which does not need composite systems or spacelike separation. Contextuality is proven either by a logical contradiction between the noncontextuality hidden variable predictions and those of quantum mechanics or by the violation of noncontextual inequality. We propose an experimental scheme of state-independent contextual inequality derived from the Mermin proof of the Kochen–Specker(KS) theorem in eight-dimensional Hilbert space, which could be observed either in an individual system or in a composite system. We also show how to resolve the compatibility problems. Our scheme can be implemented in optical systems with current experiment techniques.

Delayed Selfing in an Alpine Biennial Gentianopsis paludosa(Gentianaceae) in the Qinghai-Tibetan Plateau

Delayed selfing could provide ovules with an opportunity to be fertilized as a means of ＂pollination assurance＂ before the flowers wilt. It could, thus, be regarded as an adaptation to unpredictable pollinator environments. Within the alpine biennial Gentianopsis paludosa, the showy flowers and herkogamy at the early stage of a flower＇s life cycle may favor outcrossing. As the flower ages, anthers contact the central stigma due to the elongation of all filaments, resulting in autonomous selfing. Flower visitors are extremely rare in a high altitude population; and examination of the mating system indicates that G. paludosa is self-pollinated under natural conditions in this population. While at the lower altitude, the bumblebee visitation rate is relatively high but possibly unreliable. Stigma receptivity is the highest on the third day of anthesis, and decreases thereafter. Pollen viability is the highest when flowers open, and gradually decreases later. Self-pollination of G. paludosa occurs at the late stage of a flower＇s lifecycle when stigma receptivity and pollen viability have both decreased, suggesting delayed selfing and assurance of seed production. This delayed selfing could assure seed production under the constraints of pollinator scarcity, but ensure outcrossing when pollinators were available. Such a flexible pollination mechanism is highly adaptive in the alpine environment of the Qinghai-Tibetan Plateau.