生物技术通报 ›› 2022, Vol. 38 ›› Issue (4): 117-125.doi: 10.13560/j.cnki.biotech.bull.1985.2021-0904
• 作物品质遗传与改良专题(专题主编: 刘巧泉 教授) • 上一篇 下一篇
高蒙1(), 李富婷1, 魏湛林1, 张赛行2, 白如仟1, 尚轶1, 马玲1()
收稿日期:
2021-10-21
出版日期:
2022-04-26
发布日期:
2022-05-06
通讯作者:
马玲,女,博士,助理研究员,研究方向:分子生物学;E-mail: maling223322@126.com作者简介:
高蒙,女,硕士研究生,研究方向:生物化学与分子生物学;E-mail: simone_gao@126.com基金资助:
GAO Meng1(), LI Fu-ting1, WEI Zhan-lin1, ZHANG Sai-hang2, BAI Ru-qian1, SHANG Yi1, MA Ling1()
Received:
2021-10-21
Published:
2022-04-26
Online:
2022-05-06
摘要:
自交不亲和(self-incompatibility,SI)广泛存在于开花植物中,是防止近交促进杂交的一种途径,可有效维持物种的基因型多样性。车前科、茄科、蔷薇科等植物中的自交不亲和类型属于典型的配子体型自交不亲和(gametophytic self-incompatibility,GSI)。在GSI植物的杂交过程中,花粉管中的SCFSLF复合体(Skp1/Cullin1/F-box)作为E3泛素连接酶,参与了花粉对异我型S-核糖核酸酶(S-RNase)的降解过程,是维持杂交亲和的关键因子。本研究通过整理GSI植物SCFSLF复合体的研究进展,借助同源比对、进化树及蛋白序列分析等生物信息学分析手段,对二倍体马铃薯中SCFSLF复合体的各个组分进行了初步分析,并对各组分基因的表达量和蛋白互作情况进行分析,最终确定了马铃薯花粉管中的SCFSLF复合体可能是由Soltu.DM.06G001040,Soltu.DM.11G004330,Soltu.DM.06G022320以及相应的SLF蛋白组成的。
高蒙, 李富婷, 魏湛林, 张赛行, 白如仟, 尚轶, 马玲. 二倍体马铃薯中SCFSLF复合体的组分分析[J]. 生物技术通报, 2022, 38(4): 117-125.
GAO Meng, LI Fu-ting, WEI Zhan-lin, ZHANG Sai-hang, BAI Ru-qian, SHANG Yi, MA Ling. Component Analysis of SCFSLF Complex in Diploid Potato[J]. Biotechnology Bulletin, 2022, 38(4): 117-125.
引物名称 Primer | 引物序列 Sequence(5'-3') |
---|---|
4330-SSK1-F | ATGGAGTTGTGTTGCCAATC |
4330-SSK1-R | AATCTCAGGACCTTCAAATGC |
1040-CLU1-F | GTGTCATGTGCCAAGTACAA |
1040-CLU1-R | CATAACGTCTATCCTTGCCG |
QEF1a-F | GATGGTCAGACCCGTGAACATGC |
QEF1a-R | ACGATTTCATCATACCTAGCCTTGGAGT |
表1 马铃薯SSK1与Cullin1荧光定量引物
Table 1 Fluorescent quantitative primers for SSK1 and Cullin1 in potato(S. tuberosum)
引物名称 Primer | 引物序列 Sequence(5'-3') |
---|---|
4330-SSK1-F | ATGGAGTTGTGTTGCCAATC |
4330-SSK1-R | AATCTCAGGACCTTCAAATGC |
1040-CLU1-F | GTGTCATGTGCCAAGTACAA |
1040-CLU1-R | CATAACGTCTATCCTTGCCG |
QEF1a-F | GATGGTCAGACCCGTGAACATGC |
QEF1a-R | ACGATTTCATCATACCTAGCCTTGGAGT |
基因编号Gene ID | 基因位点Location | E值E value | |
---|---|---|---|
PiSSK1同源基因 PiSSK1-homologous gene | Soltu.DM.11G004330 | chr11:4294163..4295355 reverse | 1.71E-102 |
Soltu.DM.11G016290 | chr11:30776770..30778841 reverse | 2.83E-44 | |
Soltu.DM.11G004300 | chr11:4286144..4286630 forward | 9.28E-43 | |
Soltu.DM.10G017580 | chr10:47907512..47910644 forward | 9.30E-42 | |
Soltu.DM.01G051200 | chr01:87717008..87719187 forward | 2.54E-41 | |
Soltu.DM.11G004500 | chr11:4499656..4500133 reverse | 1.32E-40 | |
PiCUL1-P同源基因 PiCUL1-P-homologous genes | Soltu.DM.06G001040 | chr06:1465504..1471494 reverse | 0 |
Soltu.DM.06G035050 | chr06:58938032..58944924 reverse | 0 | |
Soltu.DM.09G023090 | chr09:58921771..58929345 reverse | 0 | |
Soltu.DM.01G022080 | chr01:60076281..60087621 forward | 0 | |
Soltu.DM.09G003030 | chr09:2388523..2390740 reverse | 0 | |
Soltu.DM.09G002940 | chr09:2349588..2351796 forward | 0 | |
Soltu.DM.01G022100 | chr01:60106453..60114114 forward | 0 | |
Soltu.DM.01G022150 | chr01:60164455..60171592 forward | 0 | |
Soltu.DM.09G003010 | chr09:2377567..2379367 reverse | 0 | |
Soltu.DM.01G022120 | chr01:60134989..60155075 forward | 0 | |
PiRbx1同源基因 PiRbx1-homologous genes | Soltu.DM.06G022320 | chr06:48782308..48785375 reverse | 1.82E-71 |
表2 二倍体马铃薯DM中SCFSLF复合体组分的同源比对
Table 2 Homologous comparison of SCFSLF complex components in diploid potato DM
基因编号Gene ID | 基因位点Location | E值E value | |
---|---|---|---|
PiSSK1同源基因 PiSSK1-homologous gene | Soltu.DM.11G004330 | chr11:4294163..4295355 reverse | 1.71E-102 |
Soltu.DM.11G016290 | chr11:30776770..30778841 reverse | 2.83E-44 | |
Soltu.DM.11G004300 | chr11:4286144..4286630 forward | 9.28E-43 | |
Soltu.DM.10G017580 | chr10:47907512..47910644 forward | 9.30E-42 | |
Soltu.DM.01G051200 | chr01:87717008..87719187 forward | 2.54E-41 | |
Soltu.DM.11G004500 | chr11:4499656..4500133 reverse | 1.32E-40 | |
PiCUL1-P同源基因 PiCUL1-P-homologous genes | Soltu.DM.06G001040 | chr06:1465504..1471494 reverse | 0 |
Soltu.DM.06G035050 | chr06:58938032..58944924 reverse | 0 | |
Soltu.DM.09G023090 | chr09:58921771..58929345 reverse | 0 | |
Soltu.DM.01G022080 | chr01:60076281..60087621 forward | 0 | |
Soltu.DM.09G003030 | chr09:2388523..2390740 reverse | 0 | |
Soltu.DM.09G002940 | chr09:2349588..2351796 forward | 0 | |
Soltu.DM.01G022100 | chr01:60106453..60114114 forward | 0 | |
Soltu.DM.01G022150 | chr01:60164455..60171592 forward | 0 | |
Soltu.DM.09G003010 | chr09:2377567..2379367 reverse | 0 | |
Soltu.DM.01G022120 | chr01:60134989..60155075 forward | 0 | |
PiRbx1同源基因 PiRbx1-homologous genes | Soltu.DM.06G022320 | chr06:48782308..48785375 reverse | 1.82E-71 |
图2 SSK1蛋白序列与表达量分析 A:GSI植物中SSK基因的系统进化树(Pb:Pyrus bretschneideri 梨;Md:Malus domestica 苹果;Pet:Prunus tenella 李;Cr:Citrus reticulata 橘;Ah:Antirrhinum金鱼草;Pi:Petunia inflata膨大矮牵牛;Ph:Petunia hybrida杂交矮牵牛;L:Lily百合;红色三角所在分支是蔷薇科和茄科发生分离的分支);B:GSI植物中SSK蛋白的序列比对;C:Soltu.DM.11G004330在E172不同部位中的表达量,以stamen中的表达量为对照组,利用2-∆∆Ct法进行相对基因表达量的计算
Fig. 2 SSK1 protein sequence and expression analysis A:Phylogenetic tree of SSK genes in GSI plants(The branch marked by the red triangle is the branch where Rosaceae and Solanaceae separated). B:Sequence alignment of SSK protein in GSI plants. C:The expression of Soltu.DM.11G004330 in different organs of E172 is calculated by 2-∆∆Ct with stamen as control
图3 Cullin1蛋白进化与表达量分析 A:GSI植物中CUL基因的系统进化树(Pi:Petunia inflata 膨大矮牵牛;Ph:Petunia hybrida 杂交矮牵牛;Cr:Citrus reticulata橘;Md:Malus domestica苹果;Pet:Prunus tenella李;Pb:Pyrus bretschneideri梨;Ah:Antirrhinum金鱼草;L:Lily百合;红色三角所在分支是蔷薇科和茄科发生分离的分支);B:Soltu.DM.06G001040在E172不同部位中的表达量,以stamen中的表达量为对照组,利用2-∆∆Ct法进行相对基因表达量的计算
Fig. 3 Cullin1 protein evolution and expression analysis A:Phylogenetic tree of CUL genes in GSI plants(The branch marked by the red triangle is the branch where Rosaceae and Solanaceae separated). B:The expression of Soltu.DM.06G001040 in different organs of E172 is calculated by 2-∆∆Ct with stamen as control
图5 Soltu.DM.11G004330蛋白与SLF蛋白的互作分析 A:Soltu.DM.11G004330与SLF互作的酵母双杂交验证图(-WLHA:四缺(-Trp/W,-Leu/L,-His/H,-adenine/Ade)培养板;100,10-1,10-2分别指菌的滴定浓度;pGBKT7-53和pGADT7-T用作positive control;pGBKT7-Lam和pGADT7-T用作negative control;4330是Soltu.DM.11G004330的缩写);B:Soltu.DM.11G004330与SLF互作的荧光素酶互补验证图(烟草叶片注射分区情况如左边所示,其中4330是Soltu.DM.11G004330的缩写)
Fig. 5 Interaction analysis of Soltu.DM.11G004330 and SLF A:Y2H of Soltu.DM.11G004330 and SLF(-WLHA,synthetic dropout media lacking tryptophan,leucine,histidine,and adenine(-Trp/W,-Leu/L,-His/H,-adenine/Ade);100,10-1,and 10-2 indicates concentration of E.coli;pGBKT7-53 and pGADT7-T are used as positive control;pGBKT7-Lam and pGADT7-T are used as negative control;4330 is abbreviated from Soltu.DM.11G004330). B:LUC complementation of Soltu.DM.11G004330 and SLF.(The left figure denotes how injections were carried out on tobacco leaves and 4330 was the abbreviation of Soltu.DM.11G004330)
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摘要 |
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