本研究对锈毛莓(Rubus reflexus Ker.)叶绿体基因组进行测序及组装,获得了全长为 156 247 bp,GC总含量为 30.72%的核苷酸序列,注释出 129 个基因,其中 16 个基因有 2 个拷贝,9 个基因有内含子;密码子使用偏好结果显示,自然选择对锈毛莓密码子使用偏好影响最大,其次是突变压力;系统发育结果显示,锈毛莓(R.reflexus)与黄脉莓(R.xanthoneurus)和高粱泡(R.lambertianus)亲缘关系较近;叶绿体基因组全长比较分析结果显示,锈毛莓与近缘物种之间的变异主要存在于基因间隔区;通过核苷酸多态性分析筛选出 6 个高变区,即rps16-trnQ-UUG、petN-psbM、trnT-UGU-trnL-UAA、petA-psbJ、rpl32-trnL-UAG及ycf1.研究结果揭示了锈毛莓叶绿体基因组特征及系统发育关系,为锈毛莓及悬钩子属的比较基因组学研究、物种鉴定及遗传改良提供参考依据,同时也为解决悬钩子属部分物种分类混乱问题提供了新的解决方案.

氨磺灵是一种三硝基苯胺类除草剂,在应用于植物多倍体的诱导中,取到了较好的诱导效果.本研究以氨磺灵为诱变剂,采用混培法对来自 3 个不同地点的滇杨材料进行多倍体诱导,探讨氨磺灵对滇杨的多倍化诱导效果,继而对诱导后的材料进行同质化处理,鉴定其倍性.结果显示:氨磺灵可诱导滇杨产生多倍体材料,变异率最高的处理组合为预培养 4 d后,在添加 50 mg/L氨磺灵的分化培养基(1/2 MS+TDZ 0.01mg/L+NAA 0.01mg/L)中培养 2 d,变异率可达 28.57%.多倍体植株与正常株形态差异明显,且染色体数目增加为 76 条,多倍体滇杨的荧光通道值约为二倍体滇杨的 2 倍,表明滇杨材料已加倍为四倍体.研究结果对滇杨乃至木本植物的多倍体育种具有重要的理论及实践意义.

Objective Means utilizing molecular markers to differentiate rapidly and accurately male and female Populus yunnanensis plants at young stages was investigated. Method Using simple sequence repeats (SSR), molecular markers were screened for those might associate with the sex of P. yunnanensis at seedling stages and before flowering. Result Fifteen pairs of SSR primers were selected from the reported literatures. One of them, coded BPCA90, showed clear bands and stable reaction in agarose gel detection. After rescreening the primer by polyacrylamide gel electrophoresis, a specific band of approximately 680 bp appeared in the PCR amplification of male P. yunnanensis. Subsequent testing on 30 plants using these SSR primers showed significantly more males than females in a ratio of 13 to 2. ConclusionShort of a visible morphological sign for the determination, the SSR primers screened by this study appeared to suffice the general purpose of differentiating the sex of P. yunnanensis at early growth stage.

为了解楚雄市国有林场樟檀林分场华山松无性系种子园中6个种源间生长性状的差异,在种子园内,随机选取96个无性系的5个重复,利用测高仪、测树围尺分别测量树高和胸径,共测量480个单株,并对不同华山松无性系优株的树高、胸径生长量进行方差分析.结果表明,华山松6个种源间树高和胸径存在显著差异;LSD多重比较结果显示,会泽种源树高平均值最高与楚雄和巍山种源存在显著差异,其他5个种源间树高差异不显著;会泽胸径平均值最高与宜良、南华、巍山、楚雄和腾冲种源差异显著,其他5个种源间胸径差异不显著,表明会泽种源的华山松在种子园内生长速度最快.研究结果可为无性系选育和木材加工提供重要理论和参考依据.

[目的]在滇杨幼苗时期和开花前利用形态差异对雌雄株进行性别鉴定极为困难,因此亟需探明对其性别进行快速鉴定的分子标记方法.[方法]以滇杨雌雄各 30株为材料,利用简单重复序列(SSR)筛选与滇杨性别相关的分子标记,并利用所获得的SSR标记对未知性别滇杨幼苗进行鉴定,分析滇杨苗期的性别分化情况.[结果]从已报道的相关文献中选出 15对与植物性别连锁的SSR引物,经试验筛选出 1对(BPCA90)在琼脂糖凝胶检测中显现清晰条带且反应稳定的SSR引物,并将这对引物经聚丙烯酰胺凝胶电泳进行复筛,经PCR扩增后在滇杨雄株中出现 1条特异性条带,大小在 680 bp左右.利用这对SSR引物对 30株滇杨幼苗进行性别鉴定,结果表明雄株明显多于雌株,且比例为 13∶2.[结论]试验筛选所得到的SSR引物可以用于滇杨早期进行性别鉴定.研究结果可作为滇杨早期性别鉴定的遗传标记,为今后滇杨在生产中的分性别利用提供技术支持.

[目的]明确云南楚雄华山松无性系种子园内不同种源的遗传多样性.[方法]采集种子园内6个种源的96份华山松无性系单株针叶,采用改良的CTAB法提取基因组DNA,并利用SRAP标记对其进行遗传多样性分析.[结果]筛选获得26对条带清晰、PCR反应稳定且多态性较高的SRAP引物组合.多态性位点百分率平均值为81.22％,Shannon多样性指数(Ⅰ)为0.394 4,Nei's基因多样性指数(H)为0.259 5,不同种源间的遗传分化系数(Gst)为0.116 9;华山松种子园内各无性系在种源间变异较小,而种源内表现出较高的遗传变异;6个种源可聚为3类,遗传变异丰富度与地理距离无显著相关性(P＞0.05).[结论]楚雄华山松无性系种子园内6个种源间的遗传多样性较低,变异主要存在于各种源内.地理距离对各无性系材料的遗传距离无明显影响.在该种子园内进行杂交育种时,应以分子标记结果为主、参考地理距离进行杂交亲本的选配工作.

PurposeTo analyze the complete chloroplast genome of Rubus lambertianus. MethodsThe chloroplast genome of R. lambertianus was sequenced by high-throughput sequencing. We analyzed the chloroplast genome structure, gene composition and codon preference after genome assembly and annotation. And the maximum likelihood method was used to establish phylogenetic trees in R. lambertianus, 48 Rubus and two Geun. ResultsThe chloroplast genome structure of R. lambertianus was a classical four-section structure with a size of 156 266 bp, and a total of GC content was 37.2%. The length of large single-copy region and small single-copy region was 85 849 and 18 855 bp, respectively. The length of reverse complementary repeat region was 25 781 bp. A total of 131 genes were annotated, including 86 protein-coding genes, 37 tRNA genes and eight rRNA genes. And a total of 33 codons in R. lambertianus chloroplast genome ended in A and U, among of these, 18 codons were identified as optimal codons. It was revealed that the codon preference of R. lambertianus chloroplast genome was more influenced by natural selection by using ENC-plot and other plotting analysis methods. The phylogenetic tree showed that R. lambertianus were most closely related to R. rufus, followed by R. cochinchinensis and R. reflexus. ConclusionThe chloroplast genome of R. lambertianus is large and conserved, and the genome is most closely related to R. rufus. The results of this study can be used to analyze the phylogenetic of chloroplast genome system in R. lambertianus and Rubus genus.

为促进滇黄精的工厂化生产,选取滇黄精(Polygonatum kingianum Coil.et Hemsl)当年生幼嫩根状茎作为外植体,研究适合滇黄精腋芽诱导、增殖、生根培养的外源生长调节剂的种类、浓度配比、培养基类型,并进行移栽试验.结果表明:滇黄精根状茎诱导分化的最适培养基配方为1/2MS+6-BA 2.0 mg·L-1+2,4-D 0.2 mg·L-1+蔗糖30 g·L-1+琼脂5.5 g·L-1;滇黄精带芽茎段增殖最适培养基配方为MS+6-BA 2.0 mg·L-1+NAA 0.3 mg·L-1+蔗糖30 g·L-1+琼脂5.5 g·L-1;滇黄精带芽茎段生根最适培养基配方为1/2MS+NAA 0.2 mg·L-1+蔗糖30g·L-1+琼脂5.5 g·L-1;滇黄精组培苗生长最适基质配比是腐殖质土:红泥土:珍珠岩为1∶1∶1,其成活率可达100％.

为开发华山松大量的稳定性高、特异性强的SNP标记,本研究以279份华山松针叶为试验材料,利用SLAF-seq技术,测序共获得Reads数据3 169 Mb,SLAF标签12 952 676个,多态性SLAF标签为1 456 486个,SLAF多态性百分率是11.24％,平均测序深度为20.52 x,测序平均Q30为96.58％,平均GC含量为37.76％.且采用水稻'日本晴'的测序数据用于评估试验建库的准确性,结果显示本试验比对效率在97％;且水稻'日本晴'数据的酶切效率在100％,这表明本试验酶切反应也正常.最终从12 952 676个SLAF标签中检测到了 3 469 074个群体SNP标记.这为华山松分子辅助育种、遗传多样性分析和遗传图谱构建等研究提供理论参考.

Rosa lucieae is one of the famous wild ancestors of cultivated roses and plays a very important role in horticultural research, but there is still a lack of research on the R. lucieae chloroplast genome. In this study, we used the Illumina MiSeq platform for sequencing, assembly, and annotation to obtain the R. lucieae chloroplast genome sequencing information and compared genomics, selection stress analysis, and phylogenetic analysis with 12 other chloroplast genomes of Rosa. The R. lucieae cpDNA sequence has a total length of 156,504 bp, and 130 genes are annotated. The length of all 13 studied chloroplast genomes is 156,333~157,385 bp. Their gene content, gene sequence, GC content, and IR boundary structure were highly similar. Five kinds of large repeats were detected that numbered 100~116, and SSR sequences ranged from 78 to 90 bp. Four highly differentiated regions were identified, which can be used as potential genetic markers for Rosa. Selection stress analysis showed that there was significant positive selection among the 18 genes. The phylogenetic analysis of R. lucieae and R. cymose, R. maximowicziana, R. multiflora, and R. pricei showed the closest relationship. Overall, our results provide a more comprehensive understanding of the systematic genomics and comparative genomics of Rosa.

叶绿体基因组凭借稳定的结构、高度保守的序列在植物遗传结构、比较基因组学和系统发育进化等研究中发挥着重要作用.为明确南系华山松叶绿体基因组的大小、结构、基因组成及近缘种的关系和分类地位,本研究以南系华山松为研究对象,采用2×CTAB法提取南系华山松叶绿体基因组DNA,建立测序文库,进行浅层基因组测序、组装、SSR检测、序列对比、共线性分析以及系统发育树构建.结果表明,南系华山松叶绿体基因组全长116 879 bp,GC含量为38.16％.南系华山松叶绿体基因组共编码136个基因.其中,蛋白编码基因77个,tRNA基因55个,rRNA基因4个,具有2～5份拷贝的基因共有14个(均属于tRNA基因),具有1个内含子的基因共有12个.经叶绿体基因组序列比对发现,南系华山松与新疆五针松的相似性最高,达98.852％;与欧洲云杉的相似性最低,仅为57.588％.运用最大似然法基于GTR+F+R2模型构建系统发育树,可将20种松科植物分为4类,其中,南系华山松与北系华山松虽聚为一类,但两者之间的支持率处于较低水平,仅为55％.此外,南系华山松与北系华山松叶绿体基因组序列存在碱基替换和插入缺失,编码基因数量和种类存在异同,说明南北系华山松间叶绿体基因组存在明显差异.本研究可为南系华山松遗传结构和华山松群体遗传研究提供理论依据.