The primers were designed by Primer Premier 5.0 with homologous cloning method, according to the sequence of Arabidopsis plasma membrane intrinsic proteins gene (PIP, Genbank: NP_195236.1).
Online analysis showed that nucleic acid sequence of this fragment, named BnPIP-like gene, shared the highest identity (91.7%) with Arabidopsis plasma membrane intrinsic proteins (PIP, Genbank:NP_195236.1), which belongs to the aquaporin (AQP) family.
The aquaporins or MIPs (Major Intrinsic Proteins) are proteins assembling into water channels of cell membrane and facilitate osmosis for rapid bidirectional transport of water .
Von Wiren, "Tonoplast intrinsic proteins AtTIP2;1 and AtTIP2;3 facilitate NH3 transport into the vacuole," Plant Physiology, vol.
Gustavsson, "A new subfamily of major intrinsic proteins in plants," Molecular Biology and Evolution, vol.
Abbreviations: AQP, aquaporin; ar/R, aromatic-arginine selectivity filter; Arg, arginine; ATP, adenosine triphosphate; BIB, big brain; Eglp, entomoglyceroporin; Ga, billions of years ago; GIP, GlpF-like intrinsic protein; Glp, aquaglyceroporin; HGT, horizontal gene transfer; HIP, hybrid intrinsic protein; His, histidine; Leu, leucine; LHIP, Lygus hesperus integral protein-like channel; NIP, plant nodulin 26-like integral protein; NPA, asparagine-proline-alanine; NPC, asparagine-proline-cysteine; PIP, plasma membrane intrinsic protein; RPIP, Rhodnius prolixus integral protein-like channel; SIP, small basic intrinsic protein; TIP, plant tonoplast intrinsic protein; TMD, transmembrane domain; Trp, tryptophan; XIP, X intrinsic protein; WGD, whole genome duplication.
The N-terminus is also associated with gating of fungal Aqyl (Fischer et al., 2009), while acidic pH and the [Ca.sup.2+] concentration in the cytoplasm is known to gate both plant plasma membrane intrinsic proteins (PIPs) (Tournaire-Roux et al., 2003; Alieva et al., 2006; Verdoucq et al., 2008; Frick et al., 2013) and vertebrate aquaporins (Zeuthen and Klaerke, 1999; Nemeth-Cahalan and Hall, 2000; Virkki et al., 2001; Nemeth-Cahalan et al., 2004; Chauvigne et al., 2015a).
Plant type-1 PIPs (PIP1) and type-6 nodulin intrinsic proteins (NIP6) also display little or no water transport activity when expressed in amphibian oocytes (Chaumont et al., 2000; Dordas et al., 2000; Dixit et al., 2001; Bots et al., 2005; Temmei et al., 2005; Wallace et al., 2005; Secchi et al., 2007).
Abbreviations: AQP, aquaporin; Bas, basopinacocytes; Ch, choancythes; CrAQP, aquaporin isolated in Chondrosia reniformis; DAPI, nuclear fluorescent dye 4',6-diamidino-2-phenylindole; ESEM, environmental scanning electron microscope; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; GLPs, glycerol uptake facilitators or aquaglyceroporins; MIPs, major intrinsic proteins
; NIPs, Nodulin 26-like intrinsic proteins
; NPA, highly conserved signature of Asn-Pro-Ala; Pin, pinacocytes; PIPs.