Creation and Identification of Cucumis sativus-C. hystrix Alien Addition Lines

doi:10.16420/j.issn.0513-353x.2021-0187

Acta Horticulturae Sinica ›› 2021, Vol. 48 ›› Issue (7): 1349-1358.doi: 10.16420/j.issn.0513-353x.2021-0187

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Creation and Identification of Cucumis sativus-C. hystrix Alien Addition Lines

HU Wei, LIU Yuxi, ZHAO Qinzheng, CHEN Jinfeng, LOU Qunfeng^*()

State Key Laboratory of Crop Genetics and Germplasm Enhancement,College of Horticulture,Nanjing Agricultural University,Nanjing 210095,China

Received:2021-02-22 Revised:2021-05-18 Online:2021-07-25 Published:2021-08-10
Contact: LOU Qunfeng E-mail:qflou@njau.edu.cn

Abstract

Abstract:

Alien addition lines are important interspecific materials,which can be used for introgression of interspecific genetic material and gene mapping. Creating a variety of alien addition lines plays an important role in cultivar improvement and related research of cucumber. In this experiment,based on the allotetraploid Cucumis hytivus from the cross of C. sativus and C. hystrix,two backcrosses with diploid cultivar C. sativus were carried out to obtain 22 plants of BC₂. The plant containing alien chromosome in BC₂ population was identified by GISH technique. Further,oligo-FISH technology and 12 C. hystrix chromosome specific single-copy sequence markers were used to accurately identify the alien chromosomes. Finally,a total of four different alien additional lines were screened,including three monosomic alien addition lines（MAALs）which contained C. hystrix chromosomes H06,H10,and H12,respectively,and one double alien monosomic addition line containing H06 and H10.

Key words: Cucumis sativus, Cucumis hystrix, alien addition line, oligo-FISH, single-copy sequence marker

CLC Number:

S642.2

HU Wei, LIU Yuxi, ZHAO Qinzheng, CHEN Jinfeng, LOU Qunfeng. Creation and Identification of Cucumis sativus-C. hystrix Alien Addition Lines[J]. Acta Horticulturae Sinica, 2021, 48(7): 1349-1358.

Figures/Tables 9

Fig. 1 The process of creating materials

Table 1 Corresponding relationship between cucumber oligo probe and Cucumis hystrix chromosome

黄瓜oligo探针 Cucumber oligo probe	酸黄瓜染色体 Cucumis hystrix chromosomes	物理位置/Mb Physical positions
Oligo C7-a	H01	0 ~ 19
Oligo C1-a	H02	0 ~ 4/12.5 ~ 20/24 ~ 33
Oligo C2-a	H03	12.4 ~ 22.6
Oligo C3-a	H04	25.2 ~ 40.8
Oligo C2-b	H05	0 ~ 12.4
Oligo C3-b	H06	0 ~ 25.2
Oligo C4-a	H07	0 ~ 10.5/19.5 ~ 23.3
Oligo C4-b	H08	10.5 ~ 19.5
Oligo C5-a	H09	0 ~ 4/11 ~ 25
Oligo C5-b	H10	4 ~ 16.5/20 ~ 31.9
Oligo C6-a	H11	0 ~ 17.8
Oligo C1-b	H12	4 ~ 12.5/20 ~ 24

Table 2 The chromosomes single-copy sequence marker primers in Cucumis hystrix

单拷贝序列标记Single-copy sequences marker	前引物（5′-3′）Primer forward	后引物（5′-3′）Primer reverse
ss H01	AGCCTTTTTCCTACACCATAA	ATTCCCTAGATCCCTTGACTG
ss H02	GTCTTTTACTCTTGGTTTTCG	TAATCGTTCCACCCATCACTA
ss H03	CAATAGAATCCATATCTTTGA	GTCATTCGTTTACTATCACAT
ss H04	GGGATTAAGCTTGTTTAGGTT	ATTTCCTTGGGATCACTACTT
ss H05	GAATAGGGAAAAGTTGCCGTT	TGATGCTGTCAAGTCGAGTCA
ss H06	CGACAAGTTTGACGTTGATAT	CTAGTTTGAACTGCGTGTATTA
ss H07	TATACCCAACCAACCGATTCC	ACTTTGAGGGCTTGTGTCTCC
ss H08	ATCGTAGAAAGGAAAGGAGAC	GATCAATTGTTGGGAGTGACT
ss H09	AGAAAATTTGGATGAAAACTAGC	ATAGAGAAAACGAACACGACT
ss H10	TACATTCCAATAGGGTGCCGT	GCCGTACTCACAGATTAAAAAA
ss H11	ACTACACTAGCGAGCAAGGGC	ACAGTAATGGTTTGGAAACGG
ss H12	GGACCCAACCTTACACCCTTA	TTCGAGTTTCAATTCCTCACC

Fig. 2 Cucumis allotriploid plant morphology and fruit and chromosome GISH analysis

Fig. 3 ‘Beijing Jietou’cucumber（a）and its chromosomes（b）and alien addition lines carrying one（c）and two（d）C. hystrix chromosome

Fig. 4 FISH identification of chromosomes in Cucumis sativus-chystrix monosomic alien addition lines CC-1,CC-4 and CC-5 Hy-gDNA：C. hystrix genome DNA probe;oligo：Cucumber chromosome probe.

Fig. 5 FISH identification of chromosomes in Cucumis sativus-chystrix double monosomic alien addition lines CC-9 a：Plant morphology of CC-9;b-d：One alien chromosome identified as H06;e-g：One alien chromosome identified as H10. Hy-gDNA：C. hystrix genome DNA probe;oligo：Cucumber chromosome probe.

Fig. 6 Amplification results of single-copy sequence marker primers ss H01-ss H12 for Cucumis hystrix chromosome HH：C. hystrix;HCC：Allotriploid;CC：C. sativus‘Beijing Jietou’.

Fig. 7 Amplification results of C. hystrix chromosome specific single-copy sequence marker primers in Cucumis sativus-hystrix MAALs HH：C. hystrix;HCC：allotriploid;CC：C. sativus;ss H06,ss H10 and ss H12 were C. hystrix H06,H10 and H12 chromosome single-copy sequence markers.

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Creation and Identification of Cucumis sativus-C. hystrix Alien Addition Lines

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