Molecular cytogenetic identification of wheat-Thinopyrum intermedium 2A/6St substitution strain 014-459
TAO Jun,1,2, LAN Xiu-Jin,1,*1Triticeae Research Institute, Sichuan Agricultural University, Chengdu 611130, Sichuan, China 2Mianyang Academy of Agricultural Sciences, Mianyang 621023, Sichuan, China
Abstract Thinopyrum intermedium is a valuable resource for wheat genetic improvement, and numerous wheat-Th. intermedium chromosome addition and substitution lines and partial amphiploids have been developed. Zhong 4 is a wheat-Th. intermedium partial amphidiploid, which can easily hybridize with wheat and is extensively utilized for wheat improvement. The strain 014-459 was a progeny of partial amphidiploid Zhong 4 descendant of wheat and Th. intermedium with some special characteristics such as high crude protein and wet gluten content and partial F1 of common wheat cultivars and strain 014-459 were sterility while a few others were fertility despite reciprocal cross. Molecular cytogenetic identification of strain 014-459 were detected for speculated contained fragment of Th. intermedium chromosomes based on its special characters. Genomic composition of strain 014-459 was detected by FISH, GISH, and PLUG (polymerase chain reaction-based landmark unique gene) marker analysis. Combined FISH/GISH analysis confirmed that a pair of wheat 2A chromosomes in strain 014-459 of the hybrid progeny between wheat-Thinopyrum intermedium partial amphidiploid Zhong 4 and wheat was replaced by a pair of St chromosomes from Thinopyrum intermedium. PLUG marker analysis verified that this pair belonged to the sixth homologous group. The special characters of strain 014-459 might be attributable to the substitution of Th. intermedium St chromosome into wheat. The study of molecular cytogenetic identification of strain 014-459 was probably beneficial to quality improvement and utilization of 6St chromosome of Th. intermedium in wheat breeding. Keywords：GISH;FISH;substitution strain;Pseudoroegneria strigosa;cytogenetics
PLUG标记TNAC1088、TNAC1178、TNAC1383、TNAC1408、TNAC1614、TNAC1763、TNAC1903由Hu等[38]开发, 由生工生物工程(上海)股份有限公司合成。PCR程序及酶切、电泳等按照Hu等[38]所述进行, 限制性内切酶采用的是Taq I, TAKARA公司生产。
A: FISH map of strain 014-459. Oligo-pSc119.2-2 (green), oligo-pTa535-2 (red); B: GISH map of strain 014-459. DNA of Th. intermedium used as the probe, and DNA of CS was used as the blocker; C: GISH map of strain 014-459. DNA of Pseudoroegneria strigosa was used as the probe and DNA of CS was used as the blocker; D: FISH map of strain 014-459. St2-80 was used as the probe and DNA of CS was used as the blocker; E: GISH map of pollen mother cells of strain 014-459 at the meiotic metaphase. DNA of Th. intermedium was used as the probe and DNA of CS was used as the blocker.
A: FISH map of strain 014-459. Oligo-pSc119.2-2 (green), oligo-pTa535-2 (red); B: GISH map of strain 014-459. DNA of Th. intermedium used as the probe, and DNA of CS was used as the blocker; C: GISH map of strain 014-459. DNA of rye was used as the probe and DNA of Chinese Spring (CS) wheat was used as the blocker.
A: TNAC1763 amplification product (Taq I digestion), with arrows indicating St-specific bands; B: TNAC1178 amplification product (Taq I digestion), with arrows indicating a CS-specific amplification band, which was absent from 014-459 and St (Pseudoroegneria strigosa).
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